2024-03-29T07:16:07Zhttp://repository.helmholtz-hzi.de/oai/requestoai:repository.helmholtz-hzi.de:10033/85152019-08-30T11:32:38Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Panthel, Klaus
author
Jechlinger, Wolfgang
author
Matis, Alexander
author
Rohde, Manfred
author
Szostak, Michael
author
Lubitz, Werner
author
Haas, Rainer
2007-02-19T09:34:08Z
2003-01
Infection and Immunity 2003 71(1):109-116
0019-9567
12496155
10.1128/IAI.71.1.109-116.2003
http://hdl.handle.net/10033/8515
143412
en_US
Copyright © 2003, American Society for Microbiology
Generation of Helicobacter pylori Ghosts by PhiX Protein E-Mediated Inactivation and Their Evaluation as Vaccine Candidates
URL
https://hzi.openrepository.com/bitstream/10033/8515/1/Panthel%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/8515/7/Panthel%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/86142019-08-30T11:32:39Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Rinas, U
author
Hellmuth, K
author
Kang, R
author
Seeger, A
author
Schlieker, H
2007-02-20T13:10:07Z
1995-12
Applied and Environmental Microbiology 1995 61(12):4147-4151
0099-2240
8534082
http://hdl.handle.net/10033/8614
167726
en_US
Entry of Escherichia coli into stationary phase is indicated by endogenous and exogenous accumulation of nucleobases.
URL
https://hzi.openrepository.com/bitstream/10033/8614/1/Rinas%20et%20al_final.pdf
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Rinas et al_final.pdf
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https://hzi.openrepository.com/bitstream/10033/8614/7/Rinas%20et%20al_final.pdf.txt
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Rinas et al_final.pdf.txt
oai:repository.helmholtz-hzi.de:10033/86282019-08-30T11:32:40Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Zogaj, Xhavit
author
Bokranz, Werner
author
Nimtz, Manfred
author
Römling, Ute
2007-02-20T13:26:19Z
2003-07
Infection and Immunity 2003 71(7):4151-4158
0019-9567
12819107
10.1128/IAI.71.7.4151-4158.2003
http://hdl.handle.net/10033/8628
162016
en_US
Copyright © 2003, American Society for Microbiology
Production of Cellulose and Curli Fimbriae by Members of the Family Enterobacteriaceae Isolated from the Human Gastrointestinal Tract
URL
https://hzi.openrepository.com/bitstream/10033/8628/1/Zogaj%20et%20al_final.pdf
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Zogaj et al_final.pdf
URL
https://hzi.openrepository.com/bitstream/10033/8628/7/Zogaj%20et%20al_final.pdf.txt
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Zogaj et al_final.pdf.txt
oai:repository.helmholtz-hzi.de:10033/85932019-08-30T11:37:44Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Kordel, M
author
Hofmann, B
author
Schomburg, D
author
Schmid, R D
2007-02-20T12:45:24Z
1991-08
Journal of Bacteriology 1991 173(15):4836-4841
0021-9193
1856176
http://hdl.handle.net/10033/8593
208163
Images
en_US
Extracellular lipase of Pseudomonas sp. strain ATCC 21808: purification, characterization, crystallization, and preliminary X-ray diffraction data.
URL
https://hzi.openrepository.com/bitstream/10033/8593/1/kordel%20et%20al_final.pdf
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kordel et al_final.pdf
URL
https://hzi.openrepository.com/bitstream/10033/8593/7/kordel%20et%20al_final.pdf.txt
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kordel et al_final.pdf.txt
oai:repository.helmholtz-hzi.de:10033/86732019-08-30T11:32:36Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Vallejo, Luis Felipe
author
Rinas, Ursula
2007-02-20T14:43:28Z
2004-09-02
Microbial Cell Factories 2004 3:11
1475-2859
15345063
10.1186/1475-2859-3-11
http://hdl.handle.net/10033/8673
517725
en_US
Copyright © 2004 Vallejo and Rinas; licensee BioMed Central Ltd.
Strategies for the recovery of active proteins through refolding of bacterial inclusion body proteins
URL
https://hzi.openrepository.com/bitstream/10033/8673/1/vallejo%20et%20al_final.pdf
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vallejo et al_final.pdf
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vallejo et al_final.pdf.txt
oai:repository.helmholtz-hzi.de:10033/123442019-08-30T11:32:16Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
JUDELE, Roxana
author
LASCHAT, SABINE
author
BARO, ANGELIKA
author
NIMTZ, MANFRED
2007-06-15T08:23:30Z
2006-10-09
Tetrahedron 2006, 62:41
00404020
10.1016/j.tet.2006.07.087
http://hdl.handle.net/10033/12344
A series of unsubstituted and 1,4-disubstituted gallic catecholates 1, 6 and 7 as possible candidates for wedge-shaped mesogens were prepared starting from the respective benzene derivatives 2a–c and gallic esters 5a–h. The mesomorphic properties were investigated by DSC. However, only the 4,5-dinitro derivatives 1d,f–h with C8H17 and C10H21 to C12H25 alkyl side chains displayed mesophases, as evaluated by fluidity and optical anisotropy.
en_US
Gallic Esters of 4,5-Dinitrocatechol as Potential Building Blocks for Thermotropic Liquid Crystals
Article
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URL
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oai:repository.helmholtz-hzi.de:10033/123452019-08-30T11:32:13Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Heinz, Dirk W
author
Weiss, Manfred S
author
Wendt, K Ulrich
2007-06-15T11:09:24Z
2006-01-01
Chembiochem 2006, 7(1):203-8
1439-4227
16317791
10.1002/cbic.200500459
http://hdl.handle.net/10033/12345
en
Biomacromolecular interactions, assemblies and machines: a structural view.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/12345/2/Murnau_Chembiochem_final.pdf
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Murnau_Chembiochem_final.pdf
URL
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oai:repository.helmholtz-hzi.de:10033/123562019-08-30T11:31:46Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Bösch, Andreas
author
Nimtz, Manfred
author
Mischnik, Petra
2007-06-19T12:34:55Z
2007-06-19T12:34:55Z
Cellulose 2006, 13:4, 493-507
09690239,1572882X
10.1007/s10570-005-9029-9
http://hdl.handle.net/10033/12356
n/a
Mechanistic studies on cationic ring-opening polymerisation of cyclodextrin derivatives using various Lewis acids
Article
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URL
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Bösch_final.pdf
URL
https://hzi.openrepository.com/bitstream/10033/12356/8/B%c3%b6sch_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/124302019-08-30T11:30:58Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Hunke, Cornelia
author
Hirsch, Tatjana
author
Eichler, Jutta
2007-06-25T14:52:40Z
2006-08-01
Chembiochem 2006, 7(8):1258-64
1439-4227
16810654
10.1002/cbic.200500465
http://hdl.handle.net/10033/12430
The Mena EVH1 domain, a protein-interaction module involved in actin-based cell motility, recognizes proline-rich ligand motifs, which are also present in the sequence of the surface protein ActA of Listeria monocytogenes. The interaction of ActA with host Mena EVH1 enables the bacterium to actively recruit host actin in order to spread into neighboring cells. Based on the crystal structure of Mena EVH1 in complex with a polyproline peptide ligand, we have generated a range of assembled peptides presenting the Mena EVH1 fragments that make up its discontinuous binding site for proline-rich ligands. Some of these peptides were found to inhibit the interaction of Mena EVH1 with the ligand pGolemi. One of them was further characterized at the level of individual amino acid residues; this yielded information on the contribution of individual positions of the peptides to the interaction with the ligand and identified sites for future structure optimization.
en
Structure-based synthetic mimicry of discontinuous protein binding sites: inhibitors of the interaction of Mena EVH1 domain with proline-rich ligands.
Article
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oai:repository.helmholtz-hzi.de:10033/124892019-08-30T11:25:43Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Wang, X
author
Rochon, M
author
Lamprokostopoulou, A
author
Lünsdorf, H
author
Nimtz, M
author
Römling, U
2007-07-02T08:59:50Z
2006-10-01
Cell. Mol. Life Sci. 2006, 63(19-20):2352-63
1420-682X
16952050
10.1007/s00018-006-6222-4
http://hdl.handle.net/10033/12489
Commensal Escherichia coli form biofilms at body temperature by expressing the extracellular matrix components curli fimbriae and cellulose. The role of curli fimbriae and cellulose in the interaction of commensal E. coli with the intestinal epithelial cell line HT-29 was investigated. Expression of curli fimbriae by the typical commensal isolate E. coli TOB1 caused adherence and internalization of the bacteria and triggered IL-8 production in HT-29 cells. In particular, induction of IL-8 production was complex and involved curli-bound flagellin. While cellulose alone had no effect on the interaction of TOB1 with HT-29 cells, co-expression of cellulose with curli fimbriae decreased adherence to, internalization and IL-8 induction of HT-29 cells. Investigation of a panel of commensal isolates showed a partial correlation between expression of curli fimbriae and enhanced internalization and IL-8 production. In addition, a high immunostimulatory flagellin was identified. Thus, the consequences of expression of extracellular matrix components on commensal bacterial-host interactions are complex.
en
Impact of biofilm matrix components on interaction of commensal Escherichia coli with the gastrointestinal cell line HT-29.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/12489/1/R%c3%b6mling.pdf
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Römling.pdf
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oai:repository.helmholtz-hzi.de:10033/137212019-08-30T11:33:02Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
LANGER, Olaf
author
Palme, Olov
author
Wray, Victor
author
Tokuda, Harukuni
author
Lang, Sigmund
2007-09-20T09:13:47Z
2007-09-20T09:13:47Z
Process Biochemistry 2006, 41:10
00329592
10.1016/j.procbio.2006.07.036
http://hdl.handle.net/10033/13721
n/a
Production and modification of bioactive surfactants
Article
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URL
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oai:repository.helmholtz-hzi.de:10033/146312019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Seibel, Jürgen
author
Moraru, Roxana
author
Götze, Sven
author
Buchholz, Klaus
author
Na'amnieh, Shukrallah
author
Pawlowski, Alice
author
Hecht, Hans-Jürgen
2007-11-19T15:29:22Z
2006-10-16
Carbohydr. Res. 2006, 341(14):2335-49
0008-6215
16870166
10.1016/j.carres.2006.07.001
http://hdl.handle.net/10033/14631
In the present study, we have coupled detailed acceptor and donor substrate studies of the fructosyltransferase (FTF, levansucrase) (EC 2.4.1.162) from Bacillus subtilis NCIMB 11871, with a structural model of the substrate enzyme complex in order to investigate in detail the roles of the active site amino acids in the catalytic action of the enzyme and the scope and limitation of substrates. Therefore we have isolated the ftf gene, expressed in Escherichia coli, yielding a levansucrase. Consequently, detailed acceptor property effects in the fructosylation by systematic variation of glycoside acceptors with respect to the positions (2, 3, 4 and 6) of the hydroxyl groups from equatorial to axial have been studied for preparative scale production of new oligosaccharides. Such investigations provided mechanistic insights of the FTF reaction. The configuration and the presence of the C-2 and C-3 hydroxyl groups of the glucopyranoside derivatives either as substrates or acceptors have been identified to be rate limiting for the trans-fructosylation process. The rates are rationalized on the basis of the coordination of d-glycopyranoside residues in (4)C(1) conformation with a network of amino acids by Arg360, Tyr411, Glu342, Trp85, Asp247 and Arg246 stabilization of both acceptors and substrates. In addition we also describe the first FTF reaction, which catalyzes the beta-(1-->2)-fructosyl transfer to 2-OH of L-sugars (L-glucose, L-rhamnose, L-galactose, L-fucose, L-xylose) presumably in a (1)C(4) conformation. In those conformations, the L-glycopyranosides are stabilized by the same hydrogen network. Structures of the acceptor products were determined by NMR and mass spectrometry analysis.
en
Synthesis of sucrose analogues and the mechanism of action of Bacillus subtilis fructosyltransferase (levansucrase).
Article
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URL
https://hzi.openrepository.com/bitstream/10033/14631/1/Seibel_Hecht_final.pdf
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oai:repository.helmholtz-hzi.de:10033/147332019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Schulze, Jörg O
author
Schubert, Wolf-Dieter
author
Moser, Jürgen
author
Jahn, Dieter
author
Heinz, Dirk W
2007-11-26T12:54:19Z
2006-05-19
J. Mol. Biol. 2006, 358(5):1212-20
0022-2836
16564539
10.1016/j.jmb.2006.02.064
http://hdl.handle.net/10033/14733
Glutamate-1-semialdehyde 2,1-aminomutase (GSAM) is the second enzyme in the C(5) pathway of tetrapyrrole biosynthesis found in most bacteria, in archaea and in plants. It catalyzes the transamination of glutamate-1-semialdehyde to 5-aminolevulinic acid (ALA) in a pyridoxal 5'-phosphate (PLP)-dependent manner. We present the crystal structure of GSAM from the thermophilic cyanobacterium Thermosynechococcus elongatus (GSAM(Tel)) in its PLP-bound form at 2.85A resolution. GSAM(Tel) is a symmetric homodimer, whereas GSAM from Synechococcus (GSAM(Syn)) has been described as asymmetric. The symmetry of GSAM(Tel) thus challenges the previously proposed negative cooperativity between monomers of this enzyme. Furthermore, GSAM(Tel) reveals an extensive flexible region at the interface of the proposed complex of GSAM with glutamyl-tRNA reductase (GluTR), the preceding enzyme in tetrapyrrole biosynthesis. Compared to GSAM(Syn), the monomers of GSAM(Tel) are rotated away from each other along the dimerization interface by 10 degrees . The associated flexibility of GSAM may be essential for complex formation with GluTR to occur. Unexpectedly, we find that GSAM is structurally related to 5-aminolevulinate synthase (ALAS), the ALA-producing enzyme in the Shemin pathway of alpha-proteobacteria and non-plant eukaryotes. This structural relationship applies also to the corresponding subfamilies of PLP-dependent enzymes. We thus propose that the CoA-subfamily (including ALAS) and the aminotransferase subfamily II (including GSAM) are evolutionarily closely related and that ALAS may thus have evolved from GSAM.
en
Evolutionary relationship between initial enzymes of tetrapyrrole biosynthesis.
Article
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https://hzi.openrepository.com/bitstream/10033/14733/1/Schulze%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/152302019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Schulze, Jörg O
author
Masoumi, Ava
author
Nickel, Daniel
author
Jahn, Martina
author
Jahn, Dieter
author
Schubert, Wolf-Dieter
author
Heinz, Dirk W
department
Division of Structural Biology, German Research Centre for Biotechnology (GBF), Mascheroder Weg 1, D-38124 Braunschweig, Germany.
2007-12-14T15:02:09Z
2006-09-01
Crystal structure of a non-discriminating glutamyl-tRNA synthetase. 2006, 361 (5):888-97 J. Mol. Biol.
0022-2836
16876193
10.1016/j.jmb.2006.06.054
http://hdl.handle.net/10033/15230
Journal of molecular biology
Error-free protein biosynthesis is dependent on the reliable charging of each tRNA with its cognate amino acid. Many bacteria, however, lack a glutaminyl-tRNA synthetase. In these organisms, tRNA(Gln) is initially mischarged with glutamate by a non-discriminating glutamyl-tRNA synthetase (ND-GluRS). This enzyme thus charges both tRNA(Glu) and tRNA(Gln) with glutamate. Discriminating GluRS (D-GluRS), found in some bacteria and all eukaryotes, exclusively generates Glu-tRNA(Glu). Here we present the first crystal structure of a non-discriminating GluRS from Thermosynechococcus elongatus (ND-GluRS(Tel)) in complex with glutamate at a resolution of 2.45 A. Structurally, the enzyme shares the overall architecture of the discriminating GluRS from Thermus thermophilus (D-GluRS(Tth)). We confirm experimentally that GluRS(Tel) is non-discriminating and present kinetic parameters for synthesis of Glu-tRNA(Glu) and of Glu-tRNA(Gln). Anticodons of tRNA(Glu) (34C/UUC36) and tRNA(Gln) (34C/UUG36) differ only in base 36. The pyrimidine base of C36 is specifically recognized in D-GluRS(Tth) by the residue Arg358. In ND-GluRS(Tel) this arginine residue is replaced by glycine (Gly366) presumably allowing both cytosine and the bulkier purine base G36 of tRNA(Gln) to be tolerated. Most other ND-GluRS share this structural feature, leading to relaxed substrate specificity.
en
Crystal structure of a non-discriminating glutamyl-tRNA synthetase.
Article
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oai:repository.helmholtz-hzi.de:10033/157182019-08-30T11:35:39Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Galeyeva, Yana
author
Helbig, Sarah
author
Morr, Michael
author
Sasse, Florenz
author
Nimtz, Manfred
author
Laschat, Sabine
author
Baro, Angelika
department
Institut für Organische Chemie, Universität Stuttgart, Pfaffenwaldring 55, D-70569 Stuttgart.
2008-01-04T14:51:08Z
2006-08
Total synthesis and biological evaluation of (-)-pectinatone employing a methyl-branched wax ester as key building block. 2006, 3 (8):935-41 Chem. Biodivers.
1612-1880
17193325
10.1002/cbdv.200690096
http://hdl.handle.net/10033/15718
Chemistry & biodiversity
Unnatural (-)-pectinatone ((-)-3) was prepared in five steps starting from the highly methyl-branched wax ester 4, employing bromination of the ester enolate and subsequent base-induced elimination to the enoate 6 as the key step. Both (-)-3 and the amides 8b and 8c, which were isolated as by-products in the reaction sequence, displayed antimicrobial activity and cytotoxicity.
en
Total synthesis and biological evaluation of (-)-pectinatone employing a methyl-branched wax ester as key building block.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/15718/1/Galeyeva_final.pdf
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oai:repository.helmholtz-hzi.de:10033/157372019-08-30T11:33:03Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
el-Enshasy, Hesham
author
Kleine, Joachim
author
Rinas, Ursula
department
Helmholtz zENTRUM FÜR iNFEKTIONSFORSCHUNG
2008-01-07T11:44:27Z
2006-10
00329592
10.1016/j.procbio.2006.05.024
http://hdl.handle.net/10033/15737
Process Biochemistry8(2006); 41; pp.2103-2112
Agitation effects on morphology and protein productive fractions of filamentous and pelleted growth forms of recombinant Aspergillus niger
Article
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URL
https://hzi.openrepository.com/bitstream/10033/15737/1/El-Enshasy%20et%20al_final.pdf
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URL
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oai:repository.helmholtz-hzi.de:10033/158362019-08-30T11:25:11Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Gueorguieva, Ludmila
author
Vallejo, Luis Felipe
author
Rinas, Ursula
author
Seidel-Morgenstern, Andreas
department
Otto-von-Guericke-Universität Magdeburg, Institut für Verfahrenstechnik, PO Box 4120, D-39106 Magdeburg, Germany.
2008-01-08T15:15:34Z
2006-12-01
Discontinuous and continuous separation of the monomeric and dimeric forms of human bone morphogenetic protein-2 from renaturation batches. 2006, 1135 (2):142-50notJ Chromatogr A
0021-9673
17064713
10.1016/j.chroma.2006.08.061
http://hdl.handle.net/10033/15836
Journal of chromatography. A
Bone morphogenetic protein-2 (BMP-2) is one of the most interesting of the approximately 14 BMPs which belong to the transforming-growth-factor-beta (TGF-beta) superfamily. BMP-2 induces bone formation and thus plays an important role as a pharmaceutical protein. Recently, rhBMP-2 has been produced in form of inactive inclusion bodies in Escherichia coli. After solubilization and renaturation the biologically active dimeric form of rhBMP-2 can be generated. However, inactive monomers of BMP-2 are also formed during the renaturation process which must be separated from the active dimeric BMP-2. The purpose of this paper is to present: (a) results of an experimental study of a chromatographic separation of the monomeric and dimeric forms; and (b) a concept for a continuous counter-current simulated moving bed (SMB) process. The capacity of heparin as stationary phase was estimated for different salt concentrations in the mobile phase. A simulation study of a three-zone SMB process was performed applying a two step salt gradient. The results reveal the potential of the process for the purification of the dimeric BMP-2.
en
Discontinuous and continuous separation of the monomeric and dimeric forms of human bone morphogenetic protein-2 from renaturation batches.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/15836/1/Gueorguieva%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/15836/5/Gueorguieva%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/173322019-08-30T11:33:01Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Jerz, Gerold
author
Arrey, Tabiwang N.
author
Wray, Victor
author
Du, Qizhen
author
Winterhalter, Peter
department
Helmholtz Zentrum für Infektionsforschung GmbH
2008-02-01T08:32:13Z
2007
14668564
10.1016/j.ifset.2007.03.024
http://hdl.handle.net/10033/17332
Innovative Food Science & Emerging Technologies
Structural characterization of 13(2)-hydroxy-(13(2)-S)-phaeophytin-a from leaves and stems of Amaranthus tricolor isolated by high-speed countercurrent chromatography
Article
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URL
https://hzi.openrepository.com/bitstream/10033/17332/1/Jerz%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/188252019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Zieseniss, Anke
author
Schroeder, Ulrich
author
Buchmeier, Sabine
author
Schoenenberger, Cora-Ann
author
van den Heuvel, Joop
author
Jockusch, Brigitte M
author
Illenberger, Susanne
department
Cell Biology, Zoological Institute, Technical University of Braunschweig, Biocentre, Spielmannstrasse 7, 38092 Braunschweig, Germany.
2008-02-21T13:41:29Z
2007-03
Raver1 is an integral component of muscle contractile elements. 2007, 327 (3):583-94 Cell Tissue Res.
0302-766X
17096167
10.1007/s00441-006-0322-1
http://hdl.handle.net/10033/18825
Cell and tissue research
Raver1, a ubiquitously expressed protein, was originally identified as a ligand for metavinculin, the muscle-specific isoform of the microfilament-associated protein vinculin. The protein resides primarily in the nucleus, where it colocalises and may interact with polypyrimidine-tract-binding protein, which is involved in alternative splicing processes. During skeletal muscle differentiation, raver1 translocates to the cytoplasm and eventually targets the Z-line of sarcomeres. Here, it colocalises with metavinculin, vinculin and alpha-actinin, all of which have biochemically been identified as raver1 ligands. To obtain more information about the potential role of raver1 in muscle structure and function, we have investigated its distribution and fine localisation in mouse striated and smooth muscle, by using three monoclonal antibodies that recognise epitopes in different regions of the raver1 protein. Our immunofluorescence and immunoelectron-microscopic results indicate that the cytoplasmic accumulation of raver1 is not confined to skeletal muscle but also occurs in heart and smooth muscle. Unlike vinculin and metavinculin, cytoplasmic raver1 is not restricted to costameres but additionally represents an integral part of the sarcomere. In isolated myofibrils and in ultrathin sections of skeletal muscle, raver1 has been found concentrated at the I-Z-I band. A minor fraction of raver1 is present in the nuclei of all three types of muscle. These data indicate that, during muscle differentiation, raver1 might link gene expression with structural functions of the contractile machinery of muscle.
en
Raver1 is an integral component of muscle contractile elements.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/18825/1/Zieseniss%20et%20al_final.pdf
File
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URL
https://hzi.openrepository.com/bitstream/10033/18825/8/Zieseniss%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/196732019-08-30T11:30:32Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Ganzlin, Markus
author
Marose, Stefan
author
Lu, Xin
author
Hitzmann, Bernd
author
Scheper, Thomas
author
Rinas, Ursula
department
Helmholtz Centre for Infection Research (former German Research Centre for Biotechnology - GBF), Inhoffenstr. 7, 38124 Braunschweig, Germany.
2008-03-04T14:26:26Z
2007-12-01
In situ multi-wavelength fluorescence spectroscopy as effective tool to simultaneously monitor spore germination, metabolic activity and quantitative protein production in recombinant Aspergillus niger fed-batch cultures. 2007, 132 (4):461-8 J. Biotechnol.
0168-1656
17905460
10.1016/j.jbiotec.2007.08.032
http://hdl.handle.net/10033/19673
Journal of biotechnology
The production of a mutant green fluorescent protein (S65TGFP), controlled by the maltose inducible glucoamylase promoter, was followed in situ in fed-batch cultures of recombinant Aspergillus niger using multi-wavelength fluorescence spectroscopy. Disturbance of quantitative product analysis by interfering fluorescence signals was resolved by using a set of defined combinations of excitation and emission wavelengths (lambda(ex)/lambda(em)). This technique resulted in excellent linearity between on-line signal and off-line determined S65TGFP concentrations. Spore germination was detectable in situ by monitoring the back scattered light intensity. Moreover, flavin-like fluorophores were identified as the dominating fungal host fluorophores. The time-dependent intensity of this fluorophore, potentially fungal flavin-containing oxidoreductase(s), did not correlate with the biomass concentration but correlated well with the fungal metabolic activity (e.g. respiratory activity). Other fluorophores commonly found in microbial cultures such NADH, pyridoxine and the aromatic amino acids, tryptophan, phenylalanine and tyrosine did not contribute significantly to the culture fluorescence of A. niger. Thus, multi-wavelength fluorescence spectroscopy has proven to be an effective tool for simultaneous on-line monitoring of the most relevant process variables in fungal cultures, e.g. spore germination, metabolic activity, and quantitative product formation.
en
In situ multi-wavelength fluorescence spectroscopy as effective tool to simultaneously monitor spore germination, metabolic activity and quantitative protein production in recombinant Aspergillus niger fed-batch cultures.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/19673/1/Ganzlin%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/19673/8/Ganzlin%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/196742019-08-30T11:37:44Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Rinas, Ursula
author
Hoffmann, Frank
author
Betiku, Eriola
author
Estapé, David
author
Marten, Sabine
department
Biochemical Engineering Division, GBF German Research Center for Biotechnology, Mascheroder Weg 1, D-38124 Braunschweig, Germany. URI@gbf.de
2008-03-04T14:38:17Z
2007-01-01
Inclusion body anatomy and functioning of chaperone-mediated in vivo inclusion body disassembly during high-level recombinant protein production in Escherichia coli. 2007, 127 (2):244-57 J. Biotechnol.
0168-1656
16945443
10.1016/j.jbiotec.2006.07.004
http://hdl.handle.net/10033/19674
Journal of biotechnology
During production in recombinant Escherichia coli, the human basic fibroblast growth factor (hFGF-2) partly aggregates into stable cytoplasmic inclusion bodies. These inclusion bodies additionally contain significant amounts of the heat-shock chaperone DnaK, and putative DnaK substrates such as the elongation factor Tu (ET-Tu) and the metabolic enzymes dihydrolipoamide dehydrogenase (LpdA), tryptophanase (TnaA), and d-tagatose-1,6-bisphosphate aldolase (GatY). Guanidinium hydrochloride induced disaggregation studies carried out in vitro on artificial aggregates generated through thermal aggregation of purified hFGF-2 revealed identical disaggregation profiles as hFGF-2 inclusion bodies indicating that the heterogenic composition of inclusion bodies did not influence the strength of interactions of hFGF-2 in aggregates formed in vivo as inclusion bodies compared to those generated in vitro from native and pure hFGF-2 through thermal aggregation. Compared to unfolding of native hFGF-2, higher concentrations of denaturant were required to dissolve hFGF-2 aggregates showing that more energy is required for disruption of interactions in both types of protein aggregates compared to the unfolding of the native protein. In vivo dissolution of hFGF-2 inclusion bodies was studied through coexpression of chaperones of the DnaK and GroEL family and ClpB and combinations thereof. None of the chaperone combinations was able to completely prevent the initial formation of inclusion bodies, but upon prolonged incubation mediated disaggregation of otherwise stable inclusion bodies. The GroEL system was particularly efficient in inclusion body dissolution but did not lead to a corresponding increase in soluble hFGF-2 rather was promoting the proteolysis of the recombinant growth factor. Coproduction of the disaggregating DnaK system and ClpB in conjunction with small amounts of the chaperonins GroELS was most efficient in disaggregation with concomitant formation of soluble hFGF-2. Thus, fine-balanced coproduction of chaperone combinations can play an important role in the production of soluble recombinant proteins with a high aggregation propensity not through prevention of aggregation but predominantly through their disaggregating properties.
en
Inclusion body anatomy and functioning of chaperone-mediated in vivo inclusion body disassembly during high-level recombinant protein production in Escherichia coli.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/19674/1/Rinas_final.pdf
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oai:repository.helmholtz-hzi.de:10033/197582019-08-30T11:37:23Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Hagelueken, Gregor
author
Wiehlmann, Lutz
author
Adams, Thorsten M
author
Kolmar, Harald
author
Heinz, Dirk W
author
Tümmler, Burkhard
author
Schubert, Wolf-Dieter
department
Molecular Host-Pathogen Interactions, Division of Structural Biology, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany.
2008-03-05T10:23:42Z
2007-07-24
Crystal structure of the electron transfer complex rubredoxin rubredoxin reductase of Pseudomonas aeruginosa. 2007, 104 (30):12276-81 Proc. Natl. Acad. Sci. U.S.A.
0027-8424
17636129
10.1073/pnas.0702919104
http://hdl.handle.net/10033/19758
Proceedings of the National Academy of Sciences of the United States of America
Crude oil spills represent a major ecological threat because of the chemical inertness of the constituent n-alkanes. The Gram-negative bacterium Pseudomonas aeruginosa is one of the few bacterial species able to metabolize such compounds. Three chromosomal genes, rubB, rubA1, and rubA2 coding for an NAD(P)H:rubredoxin reductase (RdxR) and two rubredoxins (Rdxs) are indispensable for this ability. They constitute an electron transport (ET) pathway that shuttles reducing equivalents from carbon metabolism to the membrane-bound alkane hydroxylases AlkB1 and AlkB2. The RdxR-Rdx system also is crucial as part of the oxidative stress response in archaea or anaerobic bacteria. The redox couple has been analyzed in detail as a model system for ET processes. We have solved the structure of RdxR of P. aeruginosa both alone and in complex with Rdx, without the need for cross-linking, and both structures were refined at 2.40- and 2.45-A resolution, respectively. RdxR consists of two cofactor-binding domains and a C-terminal domain essential for the specific recognition of Rdx. Only a small number of direct interactions govern mutual recognition of RdxR and Rdx, corroborating the transient nature of the complex. The shortest distance between the redox centers is observed to be 6.2 A.
en
Crystal structure of the electron transfer complex rubredoxin rubredoxin reductase of Pseudomonas aeruginosa.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/19758/1/Hagel%c3%bcken%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/19758/8/Hagel%c3%bcken%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/197742019-08-30T11:37:23Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Wollert, Thomas
author
Heinz, Dirk W
author
Schubert, Wolf-Dieter
department
Molecular Host-Pathogen Interactions, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany.
2008-03-05T10:30:35Z
2007-08-28
Thermodynamically reengineering the listerial invasion complex InlA/E-cadherin. 2007, 104 (35):13960-5 Proc. Natl. Acad. Sci. U.S.A.
0027-8424
17715295
10.1073/pnas.0702199104
http://hdl.handle.net/10033/19774
Proceedings of the National Academy of Sciences of the United States of America
Biological processes essentially all depend on the specific recognition between macromolecules and their interaction partners. Although many such interactions have been characterized both structurally and biophysically, the thermodynamic effects of small atomic changes remain poorly understood. Based on the crystal structure of the bacterial invasion protein internalin (InlA) of Listeria monocytogenes in complex with its human receptor E-cadherin (hEC1), we analyzed the interface to identify single amino acid substitutions in InlA that would potentially improve the overall quality of interaction and hence increase the weak binding affinity of the complex. Dissociation constants of InlA-variant/hEC1 complexes, as well as enthalpy and entropy of binding, were quantified by isothermal titration calorimetry. All single substitutions indeed significantly increase binding affinity. Structural changes were verified crystallographically at < or =2.0-A resolution, allowing thermodynamic characteristics of single substitutions to be rationalized structurally and providing unique insights into atomic contributions to binding enthalpy and entropy. Structural and thermodynamic data of all combinations of individual substitutions result in a thermodynamic network, allowing the source of cooperativity between distant recognition sites to be identified. One such pair of single substitutions improves affinity 5,000-fold. We thus demonstrate that rational reengineering of protein complexes is possible by making use of physically distant hot spots of recognition.
en
Thermodynamically reengineering the listerial invasion complex InlA/E-cadherin.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/19774/1/Wollert%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/19774/8/Wollert%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/229722019-08-30T11:30:58Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Büttner, Carina R
author
Sorg, Isabel
author
Cornelis, Guy R
author
Heinz, Dirk W
author
Niemann, Hartmut H
department
Division of Structural Biology, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany.
2008-04-11T08:00:00Z
2008-01-25
Structure of the Yersinia enterocolitica type III secretion translocator chaperone SycD. 2008, 375 (4):997-1012 J. Mol. Biol.
1089-8638
18054956
10.1016/j.jmb.2007.11.009
http://hdl.handle.net/10033/22972
Journal of molecular biology
Many Gram-negative bacteria use a type III secretion (T3S) system to directly inject effector molecules into eucaryotic cells in order to establish a symbiotic or pathogenic relationship with their host. The translocation of many T3S proteins requires specialized chaperones from the bacterial cytosol. SycD belongs to a class of T3S chaperones that assists the secretion of pore-forming translocators and, specifically chaperones the translocators YopB and YopD from enteropathogenic Yersinia enterocolitica. In addition, SycD is involved in the regulation of virulence factor biosynthesis and secretion. In this study, we present two crystal structures of Y. enterocolitica SycD at 1.95 and 2.6 A resolution, the first experimental structures of a T3S class II chaperone specific for translocators. The fold of SycD is entirely alpha-helical and reveals three tetratricopeptide repeat-like motifs that had been predicted from amino acid sequence. In both structures, SycD forms dimers utilizing residues from the first tetratricopeptide repeat motif. Using site-directed mutagenesis and size exclusion chromatography, we verified that SycD forms head-to-head homodimers in solution. Although in both structures, dimerization largely depends on the same residues, the two assemblies represent alternative dimers that exhibit different monomer orientations and overall shape. In these two distinct head-to-head dimers, both the concave and the convex surface of each monomer are accessible for interactions with the SycD binding partners YopB and YopD. A SycD variant carrying two point mutations in the dimerization interface is properly folded but defective in dimerization. Expression of this stable SycD monomer in Yersinia does not rescue the phenotype of a sycD null mutant, suggesting a physiological relevance of the dimerization interface.
en
Structure of the Yersinia enterocolitica type III secretion translocator chaperone SycD.
Article
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oai:repository.helmholtz-hzi.de:10033/234932019-08-30T11:26:07Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Wendt, K Ulrich
author
Weiss, Manfred S
author
Cramer, Patrick
author
Heinz, Dirk W
department
Department of Chemical and Analytical Sciences at Sanofi-Aventis, D-65926 Frankfurt, Germany.
2008-04-15T14:06:28Z
2008-02
Structures and diseases. 2008, 15 (2):117-20 Nat. Struct. Mol. Biol.
1545-9985
18250627
10.1038/nsmb0208-117
http://hdl.handle.net/10033/23493
Nature structural & molecular biology
en
Structures and diseases.
Article
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oai:repository.helmholtz-hzi.de:10033/239922019-08-30T11:28:24Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Niemann, Hartmut H
author
Petoukhov, Maxim V
author
Härtlein, Michael
author
Moulin, Martine
author
Gherardi, Ermanno
author
Timmins, Peter
author
Heinz, Dirk W
author
Svergun, Dmitri I
department
Division of Structural Biology, Helmholtz Center for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany.
2008-04-22T11:00:11Z
2008-03-21
X-ray and neutron small-angle scattering analysis of the complex formed by the Met receptor and the Listeria monocytogenes invasion protein InlB. 2008, 377 (2):489-500 J. Mol. Biol.
1089-8638
18262542
10.1016/j.jmb.2008.01.027
http://hdl.handle.net/10033/23992
Journal of molecular biology
The Listeria monocytogenes surface protein InlB binds to the extracellular domain of the human receptor tyrosine kinase Met, the product of the c-met proto-oncogene. InlB binding activates the Met receptor, leading to uptake of Listeria into normally nonphagocytic host cells. The N-terminal half of InlB (InlB(321)) is sufficient for Met binding and activation. The complex between this Met-binding domain of InlB and various constructs of the Met ectodomain was characterized by size exclusion chromatography and dynamic light scattering, and structural models were built using small-angle X-ray scattering and small-angle neutron scattering. Although most receptor tyrosine kinase ligands induce receptor dimerization, InlB(321) consistently binds the Met ectodomain with a 1:1 stoichiometry. A construct comprising the Sema and PSI domains of Met, although sufficient to bind the physiological Met ligand hepatocyte growth factor/scatter factor, does not form a complex with InlB(321) in solution, highlighting the importance of Met Ig domains for InlB binding. Small-angle X-ray scattering and small-angle neutron scattering measurements of ligand and receptor, both free and in complex, reveal an elongated shape for the receptor. The four Ig domains form a bent, rather than a fully extended, conformation, and InlB(321) binds to Sema and the first Ig domain of Met, in agreement with the recent crystal structure of a smaller Met fragment in complex with InlB(321). These results call into question whether receptor dimerization is the basic underlying event in InlB(321)-mediated Met activation and demonstrate differences in the mechanisms by which the physiological ligand hepatocyte growth factor/scatter factor and InlB(321) bind and activate the Met receptor.
en
X-ray and neutron small-angle scattering analysis of the complex formed by the Met receptor and the Listeria monocytogenes invasion protein InlB.
Article
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oai:repository.helmholtz-hzi.de:10033/280522019-08-30T11:26:42Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Lüer, Corinna
author
Schauer, Stefan
author
Virus, Simone
author
Schubert, Wolf-Dieter
author
Heinz, Dirk W
author
Moser, Jürgen
author
Jahn, Dieter
department
Institute of Microbiology, Technical University Braunschweig, Germany.
2008-05-26T10:42:47Z
2007-09
Glutamate recognition and hydride transfer by Escherichia coli glutamyl-tRNA reductase. 2007, 274 (17):4609-14 FEBS J.
1742-464X
17697121
10.1111/j.1742-4658.2007.05989.x
http://hdl.handle.net/10033/28052
The FEBS journal
The initial step of tetrapyrrole biosynthesis in Escherichia coli involves the NADPH-dependent reduction by glutamyl-tRNA reductase (GluTR) of tRNA-bound glutamate to glutamate-1-semialdehyde. We evaluated the contribution of the glutamate moiety of glutamyl-tRNA to substrate specificity in vitro using a range of substrates and enzyme variants. Unexpectedly, we found that tRNA(Glu) mischarged with glutamine was a substrate for purified recombinant GluTR. Similarly unexpectedly, the substitution of amino acid residues involved in glutamate side chain binding (S109A, T49V, R52K) or in stabilizing the arginine 52 glutamate interaction (glutamate 54 and histidine 99) did not abrogate enzyme activity. Replacing glutamine 116 and glutamate 114, involved in glutamate-enzyme interaction near the aminoacyl bond to tRNA(Glu), by leucine and lysine, respectively, however, did abolish reductase activity. We thus propose that the ester bond between glutamate and tRNA(Glu) represents the crucial determinant for substrate recognition by GluTR, whereas the necessity for product release by a 'back door' exit allows for a degree of structural variability in the recognition of the amino acid moiety. Analyzing the esterase activity, which occured in the absence of NADPH, of GluTR variants using the substrate 4-nitrophenyl acetate confirmed the crucial role of cysteine 50 for thioester formation. Finally, the GluTR variant Q116L was observed to lack reductase activity whereas esterase activity was retained. Structure-based molecular modeling indicated that glutamine 116 may be crucial in positioning the nicotinamide group of NADPH to allow for productive hydride transfer to the substrate. Our data thus provide new information about the distinct function of active site residues of GluTR from E. coli.
en
Glutamate recognition and hydride transfer by Escherichia coli glutamyl-tRNA reductase.
Article
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oai:repository.helmholtz-hzi.de:10033/303232019-08-30T11:29:47Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Schultheis, Ellen
author
Dreger, Michael A
author
Nimtz, Manfred
author
Wray, Victor
author
Hempel, Dietmar C
author
Nörtemann, Bernd
department
Institute of Biochemical Engineering, Technical University of Braunschweig, Braunschweig, Germany.
2008-06-23T13:38:12Z
2008-04
Structural characterization of the exopolysaccharide PS-EDIV from Sphingomonas pituitosa strain DSM 13101. 2008, 78 (6):1017-24 Appl. Microbiol. Biotechnol.
0175-7598
18286278
10.1007/s00253-008-1383-8
http://hdl.handle.net/10033/30323
Applied microbiology and biotechnology
Members of the bacterial genus Sphingomonas are known to produce highly viscous polysaccharides in solution. The exopolysaccharide PS-EDIV was produced by Sphingomonas pituitosa strain DSM 13101, purified using centrifugation, and precipitation and its structure was elucidated by 1D and 2D NMR techniques and chemical microderivatization combined with various mass spectrometric techniques. The following repeating unit of the polysaccharide could be identified: [formula: see text]. In addition, the polysaccharide also contains acetyl and glyceryl groups whose exact positions were not determined. PS-EDIV is similar in structure to a known exopolysaccharide but differs in being the first bacterial polysaccharide in which two different glucuronic acids are combined. It caused a high viscosity of the culture broth after cultivation for 48 h, although a gelation was not observed.
en
Structural characterization of the exopolysaccharide PS-EDIV from Sphingomonas pituitosa strain DSM 13101.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/30323/1/Schultheis_final.pdf
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URL
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oai:repository.helmholtz-hzi.de:10033/361322019-08-30T11:33:05Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Ganzlin, Markus
author
Rinas, Ursula
department
Helmholtz Centre for Infection Research, (Former German Research Centre for Biotechnology, GBF), Inhoffenstrasse 7, Braunschweig, Germany.
2008-08-21T11:44:25Z
2008-06-30
In-depth analysis of the Aspergillus niger glucoamylase (glaA) promoter performance using high-throughput screening and controlled bioreactor cultivation techniques. 2008, 135 (3):266-71 J. Biotechnol.
0168-1656
18501461
10.1016/j.jbiotec.2008.04.005
http://hdl.handle.net/10033/36132
Journal of biotechnology
An in-depth characterization of the Aspergillus niger glucoamylase (glaA) promoter performance was carried out on defined medium employing multi-well high-throughput screening as well as controlled batch and fed-batch bioreactor culture techniques with GFP as a fluorescent reporter protein. A variety of metabolizable carbon substrates and non-metabolizable analogs were screened with regard to their effect on the glaA expression system. The results clearly demonstrate that only starch and its hydrolytic products, including glucose, act as inducers. However, induction of the glaA expression system through the monosaccharide glucose is significantly lower compared to starch and the higher molecular weight starch degradation products. All other 26 carbon substrates tested do not induce, or even, as in the case of the easily metabolizable monosaccharide xylose, repress glaA-promoter controlled gene expression in the presence of the inducing disaccharide maltose with an increase of repression strength by increasing xylose concentrations. The complex effect of glucose on glaA-promoter controlled expression was also analyzed using non-metabolizable glucose analogs, namely 5-thio-glucose and 2-deoxyglucose, which were identified as novel and potent inducers of the glaA expression system. The results show that the induction strength depends on the inducer concentration with a maximum at defined concentrations and lower induction or even repression at concentrations above. Moreover, controlled fed-batch cultivations using a high maltose feed rate with concomitant extracellular accumulation of glucose resulted in lower levels of the reporter protein compared to cultures with a low-maltose feed rate without extracellular glucose accumulation, thus supporting the conclusion that increasing the glucose concentration beyond a critical point reduces the induction strength or may even cause repression. This way, the speed of polymer hydrolysis, glucose uptake and intracellular breakdown can be fine-tuned for optimal fungal growth and the metabolic burden for glucoamylase synthesis can be limited adequately in response to nutrient availability.
en
In-depth analysis of the Aspergillus niger glucoamylase (glaA) promoter performance using high-throughput screening and controlled bioreactor cultivation techniques.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/36132/1/Ganzlin_Rinas_final.pdf
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URL
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oai:repository.helmholtz-hzi.de:10033/477142019-08-30T11:32:16Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Gasser, Brigitte
author
Saloheimo, Markku
author
Rinas, Ursula
author
Dragosits, Martin
author
Rodríguez-Carmona, Escarlata
author
Baumann, Kristin
author
Giuliani, Maria
author
Parrilli, Ermenegilda
author
Branduardi, Paola
author
Lang, Christine
author
Porro, Danilo
author
Ferrer, Pau
author
Tutino, Maria Luisa
author
Mattanovich, Diethard
author
Villaverde, Antonio
department
University of Natural Resources and Applied Life Sciences Vienna, Department of Biotechnology, Vienna, Austria. diethard.mattanovich@boku.ac.at.
2009-01-19T15:22:12Z
2008
Protein folding and conformational stress in microbial cells producing recombinant proteins: a host comparative overview. 2008, 7:11 Microb. Cell Fact.
1475-2859
18394160
10.1186/1475-2859-7-11
http://hdl.handle.net/10033/47714
Microbial cell factories
ABSTRACT: Different species of microorganisms including yeasts, filamentous fungi and bacteria have been used in the past 25 years for the controlled production of foreign proteins of scientific, pharmacological or industrial interest. A major obstacle for protein production processes and a limit to overall success has been the abundance of misfolded polypeptides, which fail to reach their native conformation. The presence of misfolded or folding-reluctant protein species causes considerable stress in host cells. The characterization of such adverse conditions and the elicited cell responses have permitted to better understand the physiology and molecular biology of conformational stress. Therefore, microbial cell factories for recombinant protein production are depicted here as a source of knowledge that has considerably helped to picture the extremely rich landscape of in vivo protein folding, and the main cellular players of this complex process are described for the most important cell factories used for biotechnological purposes.
en
Protein folding and conformational stress in microbial cells producing recombinant proteins: a host comparative overview.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/47714/5/gasser%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/483162019-08-30T11:26:41Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Nath, Sunil
department
Structural Biology, Helmholtz Centre for Infection Research
2009-02-02T10:28:41Z
2008-09
14220067
10.3390/ijms9091784
http://hdl.handle.net/10033/48316
International Journal of Molecular Sciences
The new Unified Theory of ATP Synthesis/Hydrolysis and Muscle Contravtion, Its Manifold Fundamental Consequences and Mechanistic Implications and Its Applications in Health and Disease
Article
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URL
https://hzi.openrepository.com/bitstream/10033/48316/1/Nath_final.pdf
File
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oai:repository.helmholtz-hzi.de:10033/483212019-08-30T11:33:30Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Wiesand, Ulrich
author
Sorg, Isabel
author
Amstutz, Marlise
author
Wagner, Stefanie
author
van den Heuvel, Joop
author
Lührs, Thorsten
author
Cornelis, Guy R
author
Heinz, Dirk W
department
Division of Structural Biology, Helmholtz Centre for Infection Research (HZI), Inhoffenstrasse 7, 38124 Braunschweig, Germany.
2009-02-02T14:54:39Z
2009-01-23
Structure of the type III secretion recognition protein YscU from Yersinia enterocolitica. 2009, 385 (3):854-66 J. Mol. Biol.
1089-8638
18976663
10.1016/j.jmb.2008.10.034
http://hdl.handle.net/10033/48321
Journal of molecular biology
The inner-membrane protein YscU has an important role during the assembly of the Yersinia enterocolitica type III secretion injectisome. Its cytoplasmic domain (YscU(C)) recognizes translocators as individual substrates in the export hierarchy. Activation of YscU entails autocleavage at a conserved NPTH motif. Modification of this motif markedly changes the properties of YscU, including translocator export cessation and production of longer injectisome needles. We determined the crystal structures of the uncleaved variants N263A and N263D of YscU(C) at 2.05 A and 1.55 A resolution, respectively. The globular domain is found to consist of a central, mixed beta-sheet surrounded by alpha-helices. The NPTH motif forms a type II beta-turn connecting two beta-strands. NMR analysis of cleaved and uncleaved YscU(C) indicates that the global structure of the protein is retained in cleaved YscU(C). The structure of YscU(C) variant N263D reveals that wild type YscU(C) is poised for cleavage due to an optimal reaction geometry for nucleophilic attack of the scissile bond by the side chain of Asn263. In vivo analysis of N263Q and H266A/R314A YscU variants showed a phenotype that combines the absence of translocator secretion with normal needle-length control. Comparing the structure of YscU to those of related proteins reveals that the linker domain between the N-terminal transmembrane domain and the autocleavage domain can switch from an extended to a largely alpha-helical conformation, allowing for optimal positioning of the autocleavage domain during injectisome assembly.
en
Structure of the type III secretion recognition protein YscU from Yersinia enterocolitica.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/48321/1/Wiesand%20et%20al_final.pdf
File
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URL
https://hzi.openrepository.com/bitstream/10033/48321/7/Wiesand%20et%20al_final.pdf.txt
File
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oai:repository.helmholtz-hzi.de:10033/519732019-08-30T11:33:30Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Niemann, Hartmut H
author
Jäger, Volker
author
Butler, P Jonathan G
author
van den Heuvel, Joop
author
Schmidt, Sabine
author
Ferraris, Davide
author
Gherardi, Ermanno
author
Heinz, Dirk W
department
Division of Structural Biology, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany.
2009-03-04T10:23:57Z
2007-07-27
Structure of the human receptor tyrosine kinase met in complex with the Listeria invasion protein InlB. 2007, 130 (2):235-46 Cell
0092-8674
17662939
10.1016/j.cell.2007.05.037
http://hdl.handle.net/10033/51973
Cell
The tyrosine kinase Met, the product of the c-met proto-oncogene and the receptor for hepatocyte growth factor/scatter factor (HGF/SF), mediates signals critical for cell survival and migration. The human pathogen Listeria monocytogenes exploits Met signaling for invasion of host cells via its surface protein InlB. We present the crystal structure of the complex between a large fragment of the human Met ectodomain and the Met-binding domain of InlB. The concave face of the InlB leucine-rich repeat region interacts tightly with the first immunoglobulin-like domain of the Met stalk, a domain which does not bind HGF/SF. A second contact between InlB and the Met Sema domain locks the otherwise flexible receptor in a rigid, signaling competent conformation. Full Met activation requires the additional C-terminal domains of InlB which induce heparin-mediated receptor clustering and potent signaling. Thus, although it elicits a similar cellular response, InlB is not a structural mimic of HGF/SF.
en
Structure of the human receptor tyrosine kinase met in complex with the Listeria invasion protein InlB.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/51973/1/Nieman_final2.pdf
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oai:repository.helmholtz-hzi.de:10033/650552019-08-30T11:33:05Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Gurramkonda, Chandrasekhar
author
Adnan, Ahmad
author
Gäbel, Thomas
author
Lünsdorf, Heinrich
author
Ross, Anton
author
Nemani, Satish Kumar
author
Swaminathan, Sathyamangalam
author
Khanna, Navin
author
Rinas, Ursula
department
Helmholtz Centre for Infection Research, Braunschweig, Germany. ursula.rinas@helmholtz-hzi.de.
2009-04-16T10:59:53Z
2009
Simple high-cell density fed-batch technique for high-level recombinant protein production with Pichia pastoris: Application to intracellular production of Hepatitis B surface antigen. 2009, 8:13 Microb. Cell Fact.
1475-2859
19208244
10.1186/1475-2859-8-13
http://hdl.handle.net/10033/65055
Microbial cell factories
ABSTRACT: BACKGROUND: Hepatitis B is a serious global public health concern. Though a safe and efficacious recombinant vaccine is available, its use in several resource-poor countries is limited by cost. We have investigated the production of Hepatitis B virus surface antigen (HBsAg) using the yeast Pichia pastoris GS115 by inserting the HBsAg gene into the alcohol oxidase 1 locus. RESULTS: Large-scale production was optimized by developing a simple fed-batch process leading to enhanced product titers. Cells were first grown rapidly to high-cell density in a batch process using a simple defined medium with low salt and high glycerol concentrations. Induction of recombinant product synthesis was carried out using rather drastic conditions, namely through the addition of methanol to a final concentration of 6 g L-1. This methanol concentration was kept constant for the remainder of the cultivation through continuous methanol feeding based on the on-line signal of a flame ionization detector employed as methanol analyzer in the off-gas stream. Using this robust feeding protocol, maximum concentrations of ~7 grams HBsAg per liter culture broth were obtained. The amount of soluble HBsAg, competent for assembly into characteristic virus-like particles (VLPs), an attribute critical to its immunogenicity and efficacy as a hepatitis B vaccine, reached 2.3 grams per liter of culture broth. CONCLUSION: In comparison to the highest yields reported so far, our simple cultivation process resulted in an ~7 fold enhancement in total HBsAg production with more than 30% of soluble protein competent for assembly into VLPs. This work opens up the possibility of significantly reducing the cost of vaccine production with implications for expanding hepatitis B vaccination in resource-poor countries.
en
Simple high-cell density fed-batch technique for high-level recombinant protein production with Pichia pastoris: Application to intracellular production of Hepatitis B surface antigen.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/65055/9/gurramkonda%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/65055/15/gurramkonda%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/712332019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
de Beer, Dalene
author
Jerz, Gerold
author
Joubert, Elizabeth
author
Wray, Victor
author
Winterhalter, Peter
department
ARC Infruitec-Nietvoorbij, Stellenbosch, South Africa. dbeerd@arc.agric.za
2009-06-23T08:26:46Z
2009-05-08
Isolation of isomangiferin from honeybush (Cyclopia subternata) using high-speed counter-current chromatography and high-performance liquid chromatography. 2009, 1216 (19):4282-9 J Chromatogr A
1873-3778
19272608
10.1016/j.chroma.2009.02.056
http://hdl.handle.net/10033/71233
Journal of chromatography. A
Isomangiferin was isolated from Cyclopia subternata using a multi-step process including extraction, liquid-liquid partitioning, high-speed counter-current chromatography (HSCCC) and semi-preparative reversed-phase high-performance liquid chromatography (HPLC). Enrichment of phenolic compounds in a methanol extract of C. subternata leaves was conducted using liquid-liquid partitioning with ethyl acetate-methanol-water (1:1:2, v/v). The enriched fraction was further fractionated using HSCCC with a ternary solvent system consisting of tert-butyl methyl ether-n-butanol-acetonitrile-water (3:1:1:5, v/v). Isomangiferin was isolated by semi-preparative reversed-phase HPLC from a fraction containing mostly mangiferin and isomangiferin. The chemical structure of isomangiferin was confirmed by LC-high-resolution electrospray ionization MS, as well as one- and two-dimensional NMR spectroscopy.
en
Isolation of isomangiferin from honeybush (Cyclopia subternata) using high-speed counter-current chromatography and high-performance liquid chromatography.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/71233/1/De%20Beer%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/864412019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Kayser, Hartmut
author
Mann, Karlheinz
author
Machaidze, Gia
author
Nimtz, Manfred
author
Ringler, Philippe
author
Müller, Shirley A
author
Aebi, Ueli
department
Institut für Allgemeine Zoologie und Endokrinologie, Universität Ulm, Germany. hartmut.kayser@uni-ulm.de
2009-11-19T10:08:02Z
2009-05-29
Isolation, characterisation and molecular imaging of a high-molecular-weight insect biliprotein, a member of the hexameric arylphorin protein family. 2009, 389 (1):74-89 J. Mol. Biol.
1089-8638
19361516
10.1016/j.jmb.2009.03.075
http://hdl.handle.net/10033/86441
Journal of molecular biology
The abundant blue hemolymph protein of the last instar larvae of the moth Cerura vinula was purified and characterized by protein-analytical, spectroscopic and electron microscopic methods. Amino acid sequences obtained from a large number of cleavage peptides revealed a high level of similarity of the blue protein with arylphorins from a number of other moth species. In particular, there is a high abundance of the aromatic amino acids tyrosine and phenylalanine amounting to about 19% of total amino acids and a low content of methionine (0.8%) in the Cerura protein. The mass of the native protein complex was studied by size-exclusion chromatography, analytical ultracentrifugation, dynamic light scattering and scanning transmission electron microscopy and found to be around 500 kDa. Denaturating gel electrophoresis and mass spectrometry suggested the presence of two proteins with masses of about 85 kDa. The native Cerura protein is, therefore, a hexameric complex of two different subunits of similar size, as is known for arylphorins. The protein was further characterized as a weakly acidic (pI approximately 5.5) glycoprotein containing mannose, glucose and N-acetylglucosamine in an approximate ratio of 10:1:1. The structure proposed for the most abundant oligosaccharide of the Cerura arylphorin was the same as already identified in arylphorins from other moths. The intense blue colour of the Cerura protein is due to non-covalent association with a bilin of novel structure at an estimated protein subunit-to-ligand ratio of 3:1. Transmission electron microscopy of the biliprotein showed single particles of cylindrical shape measuring about 13 nm in diameter and 9 nm in height. A small fraction of particles of the same diameter but half the height was likely a trimeric arylphorin dissociation intermediate. Preliminary three-dimensional reconstruction based on averaged transmission electron microscopy projections of the individual particles revealed a double-trimeric structure for the hexameric Cerura biliprotein complex, suggesting it to be a dimer of trimers.
en
Isolation, characterisation and molecular imaging of a high-molecular-weight insect biliprotein, a member of the hexameric arylphorin protein family.
Article
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oai:repository.helmholtz-hzi.de:10033/929012019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Ferraris, Davide M
author
Gherardi, Ermanno
author
Di, Ying
author
Heinz, Dirk W
author
Niemann, Hartmut H
department
Division of Structural Biology, Helmholtz Centre for Infection Research (HZI), Inhoffenstrasse 7, 38124 Braunschweig, Germany. yyoshiki@riken.jp
2010-02-24T13:04:40Z
2010-01-22
Ligand-mediated dimerization of the Met Receptor tyrosine kinase by the bacterial invasion protein InlB. 2010, 395 (3):522-32 J. Mol. Biol.
1089-8638
19900460
10.1016/j.jmb.2009.10.074
http://hdl.handle.net/10033/92901
Journal of molecular biology
The Listeria monocytogenes surface protein InlB mediates bacterial invasion into host cells by activating the human receptor tyrosine kinase Met. So far, it is unknown how InlB or the physiological Met ligand hepatocyte growth factor/scatter factor causes Met dimerization, which is considered a prerequisite for receptor activation. We determined two new structures of InlB, revealing a recurring, antiparallel, dimeric arrangement, in which the two protomers interact through the convex face of the leucine-rich repeat domain. The same contact is found in one structure of the InlB-Met complex. Mutations disrupting the interprotomeric contact of InlB reduced its ability to activate Met and downstream signaling. Conversely, stabilization of this crystal contact by two intermolecular disulfide bonds generates a constitutively dimeric InlB variant with exceptionally high signaling activity, which can stimulate cell motility and cell division. These data demonstrate that the signaling-competent InlB-Met complex assembles with 2:2 stoichiometry around a back-to-back InlB dimer, enabling the direct contact between the stalk region of two Met molecules.
en
Ligand-mediated dimerization of the Met Receptor tyrosine kinase by the bacterial invasion protein InlB.
Article
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oai:repository.helmholtz-hzi.de:10033/1102362019-08-30T11:35:39Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Klink, Björn U
author
Barden, Stephan
author
Heidler, Thomas V
author
Borchers, Christina
author
Ladwein, Markus
author
Stradal, Theresia E B
author
Rottner, Klemens
author
Heinz, Dirk W
department
Division of Structural Biology, Helmholtz Zentrum für Infektionsforschung, D-38124 Braunschweig, Germany.
2010-08-24T09:31:19Z
2010-05-28
Structure of Shigella IpgB2 in complex with human RhoA: implications for the mechanism of bacterial guanine nucleotide exchange factor mimicry. 2010, 285 (22):17197-208 J. Biol. Chem.
1083-351X
20363740
10.1074/jbc.M110.107953
http://hdl.handle.net/10033/110236
The Journal of biological chemistry
A common theme in bacterial pathogenesis is the manipulation of eukaryotic cells by targeting the cytoskeleton. This is in most cases achieved either by modifying actin, or indirectly via activation of key regulators controlling actin dynamics such as Rho-GTPases. A novel group of bacterial virulence factors termed the WXXXE family has emerged as guanine nucleotide exchange factors (GEFs) for these GTPases. The precise mechanism of nucleotide exchange, however, has remained unclear. Here we report the structure of the WXXXE-protein IpgB2 from Shigella flexneri and its complex with human RhoA. We unambiguously identify IpgB2 as a bacterial RhoA-GEF and dissect the molecular mechanism of GDP release, an essential prerequisite for GTP binding. Our observations uncover that IpgB2 induces conformational changes on RhoA mimicking DbI- but not DOCK family GEFs. We also show that dissociation of the GDP.Mg(2+) complex is preceded by the displacement of the metal ion to the alpha-phosphate of the nucleotide, diminishing its affinity to the GTPase. These data refine our understanding of the mode of action not only of WXXXE GEFs but also of mammalian GEFs of the DH/PH family.
en
Structure of Shigella IpgB2 in complex with human RhoA: implications for the mechanism of bacterial guanine nucleotide exchange factor mimicry.
Article
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oai:repository.helmholtz-hzi.de:10033/1114172019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Klink, Björn U
author
Barden, Stephan
author
Heidler, Thomas V
author
Borchers, Christina
author
Ladwein, Markus
author
Stradal, Theresia E B
author
Rottner, Klemens
author
Heinz, Dirk W
department
Division of Structural Biology, Helmholtz Zentrum für Infektionsforschung, D-38124 Braunschweig, Germany.
2010-09-20T09:43:36Z
2010-05-28
Structure of Shigella IpgB2 in complex with human RhoA: implications for the mechanism of bacterial guanine nucleotide exchange factor mimicry. 2010, 285 (22):17197-208 J. Biol. Chem.
1083-351X
20363740
10.1074/jbc.M110.107953
http://hdl.handle.net/10033/111417
The Journal of biological chemistry
A common theme in bacterial pathogenesis is the manipulation of eukaryotic cells by targeting the cytoskeleton. This is in most cases achieved either by modifying actin, or indirectly via activation of key regulators controlling actin dynamics such as Rho-GTPases. A novel group of bacterial virulence factors termed the WXXXE family has emerged as guanine nucleotide exchange factors (GEFs) for these GTPases. The precise mechanism of nucleotide exchange, however, has remained unclear. Here we report the structure of the WXXXE-protein IpgB2 from Shigella flexneri and its complex with human RhoA. We unambiguously identify IpgB2 as a bacterial RhoA-GEF and dissect the molecular mechanism of GDP release, an essential prerequisite for GTP binding. Our observations uncover that IpgB2 induces conformational changes on RhoA mimicking DbI- but not DOCK family GEFs. We also show that dissociation of the GDP.Mg(2+) complex is preceded by the displacement of the metal ion to the alpha-phosphate of the nucleotide, diminishing its affinity to the GTPase. These data refine our understanding of the mode of action not only of WXXXE GEFs but also of mammalian GEFs of the DH/PH family.
en
Structure of Shigella IpgB2 in complex with human RhoA: implications for the mechanism of bacterial guanine nucleotide exchange factor mimicry.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/111417/1/Klink%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/1192102019-08-30T11:35:39Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Kämpfer, Peter
author
Busse, Hans-Jürgen
author
Tindall, Brian J
author
Nimtz, Manfred
author
Grün-Wollny, Iris
department
Institut für Angewandte Mikrobiologie, Justus-Liebig-Universität Giessen, D-35392 Giessen, Germany. peter.kaempfer@umwelt.uni-giessen.de
2011-01-12T15:07:59Z
2010-05
Nonomuraea rosea sp. nov. 2010, 60 (Pt 5):1118-24 Int. J. Syst. Evol. Microbiol.
1466-5026
19666797
10.1099/ijs.0.014845-0
http://hdl.handle.net/10033/119210
International journal of systematic and evolutionary microbiology
A Gram-positively staining, aerobic, non-motile actinomycete, strain GW 12687(T), that formed rose-pigmented colonies and branched substrate and aerial mycelia was studied in detail for its taxonomic position. On the basis of 16S rRNA gene sequence similarity studies, strain GW 12687(T) was grouped into the genus Nonomuraea, being most closely related to Nonomuraea dietziae (97.6 %), Nonomuraea africana (97.1 %), and Nonomuraea kuesteri (97.1 %). The 16S rRNA gene sequence similarity to other species of the genus Nonomuraea was < or =97 %. The chemotaxonomic characterization supported allocation of the strain to the genus Nonomuraea. The major menaquinone was MK-9(H(4)) with minor amounts of MK-9(H(2)), MK-9(H(6)), MK-9(H(0)) and MK-8(H(4)). The polar lipid profile contained the major compound diphosphatidylglycerol, moderate amounts of phosphatidylmonomethylethanolamine, phosphatidylethanolamine, hydroxy-phosphatidylethanolamine, hydroxy-phosphatidylmonomethylethanolamine, and an unknown aminophosphoglycolipid. Phosphatidylinositol mannosides and phosphatidylinositol were also present. The major fatty acids were iso- and anteiso- and 10-methyl-branched fatty acids. The results of physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain GW 12687(T) from closely related species. Thus, GW 12687(T) represents a novel species of the genus Nonomuraea, for which the name Nonomuraea rosea sp. nov. is proposed, with GW 12687(T) (=DSM 45177(T) =CCUG 56107(T)) as the type strain.
en
Nonomuraea rosea sp. nov.
Article
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URL
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oai:repository.helmholtz-hzi.de:10033/1292952019-08-30T11:37:00Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Haffke, Matthias
author
Menzel, Anja
author
Carius, Yvonne
author
Jahn, Dieter
author
Heinz, Dirk W
department
Helmholtz Zentrum für Infektionsforschung, Braunschweig, Germany.
2011-05-09T14:12:58Z
2010-09
Structures of the nucleotide-binding domain of the human ABCB6 transporter and its complexes with nucleotides. 2010, 66 (Pt 9):979-87 Acta Crystallogr. D Biol. Crystallogr.
1399-0047
20823549
10.1107/S0907444910028593
http://hdl.handle.net/10033/129295
Acta crystallographica. Section D, Biological crystallography
The human ATP-binding cassette (ABC) transporter ABCB6 is involved in haem-precursor transport across the mitochondrial membrane. The crystal structure of its nucleotide-binding domain (NBD) has been determined in the apo form and in complexes with ADP, with ADP and Mg(2+) and with ATP at high resolution. The overall structure is L-shaped and consists of two lobes, consistent with other reported NBD structures. Nucleotide binding is mediated by the highly conserved Tyr599 and the Walker A motif, and induces notable structural changes. Structural comparison with other structurally characterized NBDs and full-length ABC transporters gives the first insight into the possible catalytic mechanism of ABCB6 and the role of the N-terminal helix alpha(1) in full-length ABCB6.
en
Structures of the nucleotide-binding domain of the human ABCB6 transporter and its complexes with nucleotides.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/129295/1/Haffke%20et%20al_final..pdf
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oai:repository.helmholtz-hzi.de:10033/1358882019-08-30T11:37:23Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Darpel, Karin E
author
Langner, Kathrin F A
author
Nimtz, Manfred
author
Anthony, Simon J
author
Brownlie, Joe
author
Takamatsu, Haru-Hisa
author
Mellor, Philip S
author
Mertens, Peter P C
department
Pirbright Laboratory, Vector-borne Disease Programme, Institute for Animal Health, Woking, United Kingdom. karin.darpel@bbsrc.ac.uk
2011-07-12T14:04:34Z
2011
Saliva proteins of vector Culicoides modify structure and infectivity of bluetongue virus particles. 2011, 6 (3):e17545 PLoS ONE
1932-6203
21423801
10.1371/journal.pone.0017545
http://hdl.handle.net/10033/135888
PloS one
Bluetongue virus (BTV) and epizootic haemorrhagic disease virus (EHDV) are related orbiviruses, transmitted between their ruminant hosts primarily by certain haematophagous midge vectors (Culicoides spp.). The larger of the BTV outer-capsid proteins, 'VP2', can be cleaved by proteases (including trypsin or chymotrypsin), forming infectious subviral particles (ISVP) which have enhanced infectivity for adult Culicoides, or KC cells (a cell-line derived from C. sonorensis). We demonstrate that VP2 present on purified virus particles from 3 different BTV strains can also be cleaved by treatment with saliva from adult Culicoides. The saliva proteins from C. sonorensis (a competent BTV vector), cleaved BTV-VP2 more efficiently than those from C. nubeculosus (a less competent/non-vector species). Electrophoresis and mass spectrometry identified a trypsin-like protease in C. sonorensis saliva, which was significantly reduced or absent from C. nubeculosus saliva. Incubating purified BTV-1 with C. sonorensis saliva proteins also increased their infectivity for KC cells ∼10 fold, while infectivity for BHK cells was reduced by 2-6 fold. Treatment of an 'eastern' strain of EHDV-2 with saliva proteins of either C. sonorensis or C. nubeculosus cleaved VP2, but a 'western' strain of EHDV-2 remained unmodified. These results indicate that temperature, strain of virus and protein composition of Culicoides saliva (particularly its protease content which is dependent upon vector species), can all play a significant role in the efficiency of VP2 cleavage, influencing virus infectivity. Saliva of several other arthropod species has previously been shown to increase transmission, infectivity and virulence of certain arboviruses, by modulating and/or suppressing the mammalian immune response. The findings presented here, however, demonstrate a novel mechanism by which proteases in Culicoides saliva can also directly modify the orbivirus particle structure, leading to increased infectivity specifically for Culicoides cells and, in turn, efficiency of transmission to the insect vector.
en
Saliva proteins of vector Culicoides modify structure and infectivity of bluetongue virus particles.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/135888/1/Darpel%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/1408692019-08-30T11:37:24Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Quade, Nick
author
Dieckmann, Marieke
author
Haffke, Matthias
author
Heroven, Ann Kathrin
author
Dersch, Petra
author
Heinz, Dirk W
department
Department of Molecular Structural Biology, Helmholtz Centre for Infection Research, D-38124 Braunschweig, Germany.
2011-08-26T14:30:15Z
2011-02
Structure of the effector-binding domain of the LysR-type transcription factor RovM from Yersinia pseudotuberculosis. 2011, 67 (Pt 2):81-90 Acta Crystallogr. D Biol. Crystallogr.
1399-0047
21245528
10.1107/S0907444910049681
http://hdl.handle.net/10033/140869
Acta crystallographica. Section D, Biological crystallography
In enteropathogenic Yersinia, the expression of several early-phase virulence factors such as invasin is tightly regulated in response to environmental cues. The responsible regulatory network is complex, involving several regulatory RNAs and proteins such as the LysR-type transcription regulator (LTTR) RovM. In this study, the crystal structure of the effector-binding domain (EBD) of RovM, the first LTTR protein described as being involved in virulence regulation, was determined at a resolution of 2.4 Å. Size-exclusion chromatography and comparison with structures of full-length LTTRs show that RovM is most likely to adopt a tetrameric arrangement with two distant DNA-binding domains (DBDs), causing the DNA to bend around it. Additionally, a cavity was detected in RovM which could bind small inducer molecules.
en
Structure of the effector-binding domain of the LysR-type transcription factor RovM from Yersinia pseudotuberculosis.
Article
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URL
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URL
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oai:repository.helmholtz-hzi.de:10033/2023692019-08-30T11:31:49Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Strube, Christian P
author
Homann, Arne
author
Gamer, Martin
author
Jahn, Dieter
author
Seibel, Jürgen
author
Heinz, Dirk W
department
Department of Molecular Structural Biology, Helmholtz-Centre for Infection Research, Inhoffenstrasse 7B, 38124 Braunschweig, Germany.
2012-01-11T13:03:17Z
2011-05-20
Polysaccharide synthesis of the levansucrase SacB from Bacillus megaterium is controlled by distinct surface motifs. 2011, 286 (20):17593-600 J. Biol. Chem.
1083-351X
21454585
10.1074/jbc.M110.203166
http://hdl.handle.net/10033/202369
The Journal of biological chemistry
Despite the widespread biological function of carbohydrates, the polysaccharide synthesis mechanisms of glycosyltransferases remain largely unexplored. Bacterial levansucrases (glycoside hydrolase family 68) synthesize high molecular weight, β-(2,6)-linked levan from sucrose by transfer of fructosyl units. The kinetic and biochemical characterization of Bacillus megaterium levansucrase SacB variants Y247A, Y247W, N252A, D257A, and K373A reveal novel surface motifs remote from the sucrose binding site with distinct influence on the polysaccharide product spectrum. The wild type activity (k(cat)) and substrate affinity (K(m)) are maintained. The structures of the SacB variants reveal clearly distinguishable subsites for polysaccharide synthesis as well as an intact active site architecture. These results lead to a new understanding of polysaccharide synthesis mechanisms. The identified surface motifs are discussed in the context of related glycosyltransferases.
en
Polysaccharide synthesis of the levansucrase SacB from Bacillus megaterium is controlled by distinct surface motifs.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/202369/1/Strube%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/2087492019-08-30T11:30:58Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Busso, Didier
author
Peleg, Yoav
author
Heidebrecht, Tatjana
author
Romier, Christophe
author
Jacobovitch, Yossi
author
Dantes, Ada
author
Salim, Loubna
author
Troesch, Edouard
author
Schuetz, Anja
author
Heinemann, Udo
author
Folkers, Gert E
author
Geerlof, Arie
author
Wilmanns, Matthias
author
Polewacz, Andrea
author
Quedenau, Claudia
author
Büssow, Konrad
author
Adamson, Rachel
author
Blagova, Elena
author
Walton, Julia
author
Cartwright, Jared L
author
Bird, Louise E
author
Owens, Raymond J
author
Berrow, Nick S
author
Wilson, Keith S
author
Sussman, Joel L
author
Perrakis, Anastassis
author
Celie, Patrick H N
department
Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Institut National de Santé et de Recherche Médicale (Inserm), U964/Centre National deRecherche Scientifique (CNRS), UMR 7104, Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch, France.
2012-02-03T12:10:12Z
2011-08
Expression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study. 2011, 175 (2):159-70 J. Struct. Biol.
1095-8657
21382497
10.1016/j.jsb.2011.03.004
http://hdl.handle.net/10033/208749
Journal of structural biology
Escherichia coli (E. coli) remains the most commonly used host for recombinant protein expression. It is well known that a variety of experimental factors influence the protein production level as well as the solubility profile of over-expressed proteins. This becomes increasingly important for optimizing production of protein complexes using co-expression strategies. In this study, we focus on the effect of the choice of the expression vector system: by standardizing experimental factors including bacterial strain, cultivation temperature and growth medium composition, we compare the effectiveness of expression technologies used by the partners of the Structural Proteomics in Europe 2 (SPINE2-complexes) consortium. Four different protein complexes, including three binary and one ternary complex, all known to be produced in the soluble form in E. coli, are used as the benchmark targets. The respective genes were cloned by each partner into their preferred set of vectors. The resulting constructs were then used for comparative co-expression analysis done in parallel and under identical conditions at a single site. Our data show that multiple strategies can be applied for the expression of protein complexes in high yield. While there is no 'silver bullet' approach that was infallible even for this small test set, our observations are useful as a guideline to delineate co-expression strategies for particular protein complexes.
en
Expression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study.
Article
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oai:repository.helmholtz-hzi.de:10033/2155242019-08-30T11:25:43Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Storbeck, Sonja
author
Saha, Sayantan
author
Krausze, Joern
author
Klink, Björn U
author
Heinz, Dirk W
author
Layer, Gunhild
department
Institute of Microbiology, Technische Universität Braunschweig, 38106 Braunschweig, Germany.
2012-03-13T13:12:18Z
2011-07-29
Crystal structure of the heme d1 biosynthesis enzyme NirE in complex with its substrate reveals new insights into the catalytic mechanism of S-adenosyl-L-methionine-dependent uroporphyrinogen III methyltransferases. 2011, 286 (30):26754-67 J. Biol. Chem.
1083-351X
21632530
10.1074/jbc.M111.239855
http://hdl.handle.net/10033/215524
The Journal of biological chemistry
During the biosynthesis of heme d(1), the essential cofactor of cytochrome cd(1) nitrite reductase, the NirE protein catalyzes the methylation of uroporphyrinogen III to precorrin-2 using S-adenosyl-L-methionine (SAM) as the methyl group donor. The crystal structure of Pseudomonas aeruginosa NirE in complex with its substrate uroporphyrinogen III and the reaction by-product S-adenosyl-L-homocysteine (SAH) was solved to 2.0 Å resolution. This represents the first enzyme-substrate complex structure for a SAM-dependent uroporphyrinogen III methyltransferase. The large substrate binds on top of the SAH in a "puckered" conformation in which the two pyrrole rings facing each other point into the same direction either upward or downward. Three arginine residues, a histidine, and a methionine are involved in the coordination of uroporphyrinogen III. Through site-directed mutagenesis of the nirE gene and biochemical characterization of the corresponding NirE variants the amino acid residues Arg-111, Glu-114, and Arg-149 were identified to be involved in NirE catalysis. Based on our structural and biochemical findings, we propose a potential catalytic mechanism for NirE in which the methyl transfer reaction is initiated by an arginine catalyzed proton abstraction from the C-20 position of the substrate.
en
Archived with thanks to The Journal of biological chemistry
Crystal structure of the heme d1 biosynthesis enzyme NirE in complex with its substrate reveals new insights into the catalytic mechanism of S-adenosyl-L-methionine-dependent uroporphyrinogen III methyltransferases.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/215524/1/Storbeck%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/2266712019-08-30T11:36:32Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Wilke, Sonja
author
Groebe, Lothar
author
Maffenbeier, Vitali
author
Jäger, Volker
author
Gossen, Manfred
author
Josewski, Jörn
author
Duda, Agathe
author
Polle, Lilia
author
Owens, Raymond J
author
Wirth, Dagmar
author
Heinz, Dirk W
author
van den Heuvel, Joop
author
Büssow, Konrad
department
Department of Molecular Structural Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany.
2012-05-30T08:49:37Z
2011
Streamlining homogeneous glycoprotein production for biophysical and structural applications by targeted cell line development. 2011, 6 (12):e27829 PLoS ONE
1932-6203
22174749
10.1371/journal.pone.0027829
http://hdl.handle.net/10033/226671
PloS one
Studying the biophysical characteristics of glycosylated proteins and solving their three-dimensional structures requires homogeneous recombinant protein of high quality.We introduce here a new approach to produce glycoproteins in homogenous form with the well-established, glycosylation mutant CHO Lec3.2.8.1 cells. Using preparative cell sorting, stable, high-expressing GFP 'master' cell lines were generated that can be converted fast and reliably by targeted integration via Flp recombinase-mediated cassette exchange (RMCE) to produce any glycoprotein. Small-scale transient transfection of HEK293 cells was used to identify genetically engineered constructs suitable for constructing stable cell lines. Stable cell lines expressing 10 different proteins were established. The system was validated by expression, purification, deglycosylation and crystallization of the heavily glycosylated luminal domains of lysosome-associated membrane proteins (LAMP).
en
Archived with thanks to PloS one
Streamlining homogeneous glycoprotein production for biophysical and structural applications by targeted cell line development.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/226671/1/Wilke%20et%20al_final.pdf
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Wilke et al_final.pdf
URL
https://hzi.openrepository.com/bitstream/10033/226671/8/Wilke%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/2276762019-08-30T11:36:59Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
König, Sebastian
author
Nimtz, Manfred
author
Scheiter, Maxi
author
Ljunggren, Hans-Gustaf
author
Bryceson, Yenan T
author
Jänsch, Lothar
department
Department of Molecular Structural Biology, Helmholtz-Zentrum für Infektionsforschung, Braunschweig, Germany.
2012-06-06T14:18:27Z
2012
Kinome analysis of receptor-induced phosphorylation in human natural killer cells. 2012, 7 (1):e29672 PLoS ONE
1932-6203
22238634
10.1371/journal.pone.0029672
http://hdl.handle.net/10033/227676
PloS one
Natural killer (NK) cells contribute to the defense against infected and transformed cells through the engagement of multiple germline-encoded activation receptors. Stimulation of the Fc receptor CD16 alone is sufficient for NK cell activation, whereas other receptors, such as 2B4 (CD244) and DNAM-1 (CD226), act synergistically. After receptor engagement, protein kinases play a major role in signaling networks controlling NK cell effector functions. However, it has not been characterized systematically which of all kinases encoded by the human genome (kinome) are involved in NK cell activation.
en
Archived with thanks to PloS one
Kinome analysis of receptor-induced phosphorylation in human natural killer cells.
Article
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URL
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oai:repository.helmholtz-hzi.de:10033/2309472019-08-30T11:33:28Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Langlotz, Christine
author
Schollmeyer, Martin
author
Coplin, David L.
author
Nimtz, Manfred
author
Geider, Klaus
department
Max-Planck-Institute for Cell Biology, Ladenburg, Germany
2012-06-27T09:29:52Z
2012-06-27
Biosynthesis of the repeating units of the exopolysaccharides amylovoran from Erwinia amylovora and stewartan from Pantoea stewartii 2011, 75 (4):163 Physiological and Molecular Plant Pathology
08855765
10.1016/j.pmpp.2011.04.001
http://hdl.handle.net/10033/230947
Physiological and Molecular Plant Pathology
Archived with thanks to Physiological and Molecular Plant Pathology
Biosynthesis of the repeating units of the exopolysaccharides amylovoran from Erwinia amylovora and stewartan from Pantoea stewartii
Article
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URL
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URL
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oai:repository.helmholtz-hzi.de:10033/2451912019-08-30T11:27:46Zcom_10033_620626com_10033_6832col_10033_620627col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Wilke, Sonja
author
Krausze, Joern
author
Büssow, Konrad
department
Department of Molecular Structural Biology, Helmholtz Centre for Infection Research, Inhoffenstr, 7, 38124 Braunschweig, Germany. konrad@buessow.com.
2012-09-20T09:22:54Z
2012
Crystal structure of the conserved domain of the DC lysosomal associated membrane protein: implications for the lysosomal glycocalyx. 2012, 10:62 BMC Biol.
1741-7007
22809326
10.1186/1741-7007-10-62
http://hdl.handle.net/10033/245191
BMC biology
ABSTRACT:
en
Archived with thanks to BMC biology
Crystal structure of the conserved domain of the DC lysosomal associated membrane protein: implications for the lysosomal glycocalyx.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/245191/1/wilke%20et%20al_final.pdf
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URL
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oai:repository.helmholtz-hzi.de:10033/2509932019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
de Groot, Jens C
author
Schlüter, Kai
author
Carius, Yvonne
author
Quedenau, Claudia
author
Vingadassalom, Didier
author
Faix, Jan
author
Weiss, Stefanie M
author
Reichelt, Joachim
author
Standfuss-Gabisch, Christine
author
Lesser, Cammie F
author
Leong, John M
author
Heinz, Dirk W
author
Büssow, Konrad
author
Stradal, Theresia E B
department
Division of Structural Biology, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany.
2012-11-05T14:50:53Z
2011-09-07
Structural basis for complex formation between human IRSp53 and the translocated intimin receptor Tir of enterohemorrhagic E. coli. 2011, 19 (9):1294-306 Structure
1878-4186
21893288
10.1016/j.str.2011.06.015
http://hdl.handle.net/10033/250993
Structure (London, England : 1993)
Actin assembly beneath enterohemorrhagic E. coli (EHEC) attached to its host cell is triggered by the intracellular interaction of its translocated effector proteins Tir and EspF(U) with human IRSp53 family proteins and N-WASP. Here, we report the structure of the N-terminal I-BAR domain of IRSp53 in complex with a Tir-derived peptide, in which the homodimeric I-BAR domain binds two Tir molecules aligned in parallel. This arrangement provides a protein scaffold linking the bacterium to the host cell's actin polymerization machinery. The structure uncovers a specific peptide-binding site on the I-BAR surface, conserved between IRSp53 and IRTKS. The Tir Asn-Pro-Tyr (NPY) motif, essential for pedestal formation, is specifically recognized by this binding site. The site was confirmed by mutagenesis and in vivo-binding assays. It is possible that IRSp53 utilizes the NPY-binding site for additional interactions with as yet unknown partners within the host cell.
en
Archived with thanks to Structure (London, England : 1993)
Structural basis for complex formation between human IRSp53 and the translocated intimin receptor Tir of enterohemorrhagic E. coli.
Article
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https://hzi.openrepository.com/bitstream/10033/250993/1/deGroot%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/2707952019-08-30T11:27:46Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Glotzbach, Bernhard
author
Schmelz, Stefan
author
Reinwarth, Michael
author
Christmann, Andreas
author
Heinz, Dirk W
author
Kolmar, Harald
department
Institute for Organic Chemistry and Biochemistry, Technische Universität Darmstadt, Germany.
2013-03-01T09:44:17Z
2013-01
Structural characterization of Spinacia oleracea trypsin inhibitor III (SOTI-III). 2013, 69 (Pt 1):114-20 Acta Crystallogr. D Biol. Crystallogr.
1399-0047
23275169
10.1107/S0907444912043880
http://hdl.handle.net/10033/270795
Acta crystallographica. Section D, Biological crystallography
In recent decades, several canonical serine protease inhibitor families have been classified and characterized. In contrast to most trypsin inhibitors, those from garden four o'clock (Mirabilis jalapa) and spinach (Spinacia oleracea) do not share sequence similarity and have been proposed to form the new Mirabilis serine protease inhibitor family. These 30-40-amino-acid inhibitors possess a defined disulfide-bridge topology and belong to the cystine-knot miniproteins (knottins). To date, no atomic structure of this inhibitor family has been solved. Here, the first structure of S. oleracea trypsin inhibitor III (SOTI-III), in complex with bovine pancreatic trypsin, is reported. The inhibitor was synthesized by solid-phase peptide synthesis on a multi-milligram scale and was assayed to test its inhibitory activity and binding properties. The structure confirmed the proposed cystine-bridge topology. The structural features of SOTI-III suggest that it belongs to a new canonical serine protease inhibitor family with promising properties for use in protein-engineering and medical applications.
en
Archived with thanks to Acta crystallographica. Section D, Biological crystallography
Structural characterization of Spinacia oleracea trypsin inhibitor III (SOTI-III).
Article
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URL
https://hzi.openrepository.com/bitstream/10033/270795/1/Glotzbach%20et%20al_final.pdf
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https://hzi.openrepository.com/bitstream/10033/270795/8/Glotzbach%20et%20al_final.pdf.txt
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oai:repository.helmholtz-hzi.de:10033/2931262019-08-30T11:34:22Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Zelena, Katerina
author
Takenberg, Meike
author
Lunkenbein, Stefan
author
Woche, Susanne K
author
Nimtz, Manfred
author
Berger, Ralf G
department
Naturwissenschaftliche Fakultät der Leibniz Universität Hannover, Institut für Lebensmittelchemie, Hannover, Germany.
2013-05-31T09:30:11Z
2013-03
PfaH2: A novel hydrophobin from the ascomycete Paecilomyces farinosus. 2013, 60 (2):147-54 Biotechnol. Appl. Biochem.
1470-8744
23600571
10.1002/bab.1077
http://hdl.handle.net/10033/293126
Biotechnology and applied biochemistry
The pfah2 gene coding for a novel hydrophobin PfaH2 from the ascomycete Paecilomyces farinosus was identified during sequencing of random clones from a cDNA library. The corresponding protein sequence of PfaH2 deduced from the cDNA comprised 134 amino acids (aa). A 16 aa signal sequence preceded the N-terminus of the mature protein. PfaH2 belonged to the class Ia hydrophobins. The protein was isolated using trifluoroacetic acid extraction and purified via SDS-PAGE and high-performance liquid chromatography. The surface activity of the recently described PfaH1 and of PfaH2 was compared by the determination of contact angles (CAs) on glass slides and Teflon tape, and the CA of distilled water droplets was measured on glass slides coated with hydrophobin PfaH1 or PfaH2. Surprisingly, both hydrophobins adsorbed to hydrophilic surfaces and changed their physicochemical properties to a similar quantitative extent, although little aa sequence homology was found.
en
Archived with thanks to Biotechnology and applied biochemistry
PfaH2: A novel hydrophobin from the ascomycete Paecilomyces farinosus.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/293126/1/Zelena%20et%20al_final.pdf
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URL
https://hzi.openrepository.com/bitstream/10033/293126/11/Fig-1%20to%20Fig-4.pdf.txt
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URL
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oai:repository.helmholtz-hzi.de:10033/2972262019-08-30T11:28:46Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Kudryashev, M.
author
Stenta, M.
author
Schmelz, S.
author
Amstutz, M.
author
Wiesand, U.
author
Castano-Diez, D.
author
Degiacomi, M. T.
author
Munnich, S.
author
Bleck, C. K.
author
Kowal, J.
author
Diepold, A.
author
Heinz, D. W.
author
Dal Peraro, M.
author
Cornelis, G. R.
author
Stahlberg, H.
department
Department of Molecular Structural Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany
2013-08-01T13:33:22Z
2013-07-30
In situ structural analysis of the Yersinia enterocolitica injectisome 2013, 2 (0):e00792 eLife
2050-084X
10.7554/eLife.00792
http://hdl.handle.net/10033/297226
eLife
Archived with thanks to eLife
In situ structural analysis of the Yersinia enterocolitica injectisome
Article
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oai:repository.helmholtz-hzi.de:10033/3048122019-08-30T11:33:30Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Hofmann, Julia
author
Heider, Christine
author
Li, Wei
author
Krausze, Joern
author
Roessle, Manfred
author
Wilharm, Gottfried
department
Robert Koch-Institute, Wernigerode Branch, Burgstr. 37, D-38855 Wernigerode, Germany.
2013-10-31T12:11:40Z
2013-04
Recombinant production of Yersinia enterocolitica pyruvate kinase isoenzymes PykA and PykF. 2013, 88 (2):243-7 Protein Expr. Purif.
1096-0279
23384479
10.1016/j.pep.2013.01.010
http://hdl.handle.net/10033/304812
Protein expression and purification
The glycolytic enzyme pyruvate kinase (PK) generates ATP from ADP through substrate-level phosphorylation powered by the conversion of phosphoenolpyruvate to pyruvate. In contrast to other bacteria, Enterobacteriaceae, such as pathogenic yersiniae, harbour two pyruvate kinases encoded by pykA and pykF. The individual roles of these isoenzymes are poorly understood. In an attempt to make the Yersinia enterocolitica pyruvate kinases PykA and PykF amenable to structural and functional characterisation, we produced them untagged in Escherichia coli and purified them to near homogeneity through a combination of ion exchange and size exclusion chromatography, yielding more than 180 mg per litre of batch culture. The solution structure of PykA and PykF was analysed through small angle X-ray scattering which revealed the formation of PykA and PykF tetramers and confirmed the binding of the allosteric effector fructose-1,6-bisphosphate (FBP) to PykF but not to PykA.
en
Archived with thanks to Protein expression and purification
Recombinant production of Yersinia enterocolitica pyruvate kinase isoenzymes PykA and PykF.
Article
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URL
https://hzi.openrepository.com/bitstream/10033/304812/1/Hofmann%20et%20al_final.pdf
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oai:repository.helmholtz-hzi.de:10033/6210442018-12-20T01:42:22Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Sun, Jibin
author
Lu, Xin
author
Rinas, Ursula
author
Ping Zeng, An
2017-08-04T08:46:09Z
2007-09-04
Genome Biology. 2007 Sep 04;8(9):R182
http://dx.doi.org/10.1186/gb-2007-8-9-r182
Abstract Background Aspergillus niger is an important industrial microorganism for the production of both metabolites, such as citric acid, and proteins, such as fungal enzymes or heterologous proteins. Despite its extensive industrial applications, the genetic inventory of this fungus is only partially understood. The recently released genome sequence opens a new horizon for both scientific studies and biotechnological applications. Results Here, we present the first genome-scale metabolic network for A. niger and an in-depth genomic comparison of this species to seven other fungi to disclose its metabolic peculiarities. The raw genomic sequences of A. niger ATCC 9029 were first annotated. The reconstructed metabolic network is based on the annotation of two A. niger genomes, CBS 513.88 and ATCC 9029, including enzymes with 988 unique EC numbers, 2,443 reactions and 2,349 metabolites. More than 1,100 enzyme-coding genes are unique to A. niger in comparison to the other seven fungi. For example, we identified additional copies of genes such as those encoding alternative mitochondrial oxidoreductase and citrate synthase in A. niger, which might contribute to the high citric acid production efficiency of this species. Moreover, nine genes were identified as encoding enzymes with EC numbers exclusively found in A. niger, mostly involved in the biosynthesis of complex secondary metabolites and degradation of aromatic compounds. Conclusion The genome-level reconstruction of the metabolic network and genome-based metabolic comparison disclose peculiarities of A. niger highly relevant to its biotechnological applications and should contribute to future rational metabolic design and systems biology studies of this black mold and related species.
en
Metabolic peculiarities of Aspergillus niger disclosed by comparative metabolic genomics
Journal Article
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oai:repository.helmholtz-hzi.de:10033/5788812019-08-30T11:33:30Zcom_10033_6832com_10033_311308col_10033_620777col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Sun, Jibin
author
Lu, Xin
author
Rinas, Ursula
author
Zeng, An Ping
department
Helmholtz Centre for infection research, Inhoffenstr. 7, D-38124 Braunschweig, Germany.
2015-09-29T11:44:49Z
2007
Metabolic peculiarities of Aspergillus niger disclosed by comparative metabolic genomics. 2007, 8 (9):R182 Genome Biol.
1474-760X
17784953
10.1186/gb-2007-8-9-r182
http://hdl.handle.net/10033/578881
Genome biology
Aspergillus niger is an important industrial microorganism for the production of both metabolites, such as citric acid, and proteins, such as fungal enzymes or heterologous proteins. Despite its extensive industrial applications, the genetic inventory of this fungus is only partially understood. The recently released genome sequence opens a new horizon for both scientific studies and biotechnological applications.
en
Metabolic peculiarities of Aspergillus niger disclosed by comparative metabolic genomics.
Article
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oai:repository.helmholtz-hzi.de:10033/5823392019-08-30T11:34:48Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Hebecker, Stefanie
author
Krausze, Joern
author
Hasenkampf, Tatjana
author
Schneider, Julia
author
Groenewold, Maike
author
Reichelt, Joachim
author
Jahn, Dieter
author
Heinz, Dirk W
author
Moser, Jürgen
department
Helmholtz Centre for infection research, Inhoffenstr. 7, D-38124 Braunschweig, Germany.
2015-11-18T15:59:03Z
2015-08-25
Structures of two bacterial resistance factors mediating tRNA-dependent aminoacylation of phosphatidylglycerol with lysine or alanine. 2015, 112 (34):10691-6 Proc. Natl. Acad. Sci. U.S.A.
1091-6490
26261323
10.1073/pnas.1511167112
http://hdl.handle.net/10033/582339
Proceedings of the National Academy of Sciences of the United States of America
The cytoplasmic membrane is probably the most important physical barrier between microbes and the surrounding habitat. Aminoacylation of the polar head group of the phospholipid phosphatidylglycerol (PG) catalyzed by Ala-tRNA(Ala)-dependent alanyl-phosphatidylglycerol synthase (A-PGS) or by Lys-tRNA(Lys)-dependent lysyl-phosphatidylglycerol synthase (L-PGS) enables bacteria to cope with cationic peptides that are harmful to the integrity of the cell membrane. Accordingly, these synthases also have been designated as multiple peptide resistance factors (MprF). They consist of a separable C-terminal catalytic domain and an N-terminal transmembrane flippase domain. Here we present the X-ray crystallographic structure of the catalytic domain of A-PGS from the opportunistic human pathogen Pseudomonas aeruginosa. In parallel, the structure of the related lysyl-phosphatidylglycerol-specific L-PGS domain from Bacillus licheniformis in complex with the substrate analog L-lysine amide is presented. Both proteins reveal a continuous tunnel that allows the hydrophobic lipid substrate PG and the polar aminoacyl-tRNA substrate to access the catalytic site from opposite directions. Substrate recognition of A-PGS versus L-PGS was investigated using misacylated tRNA variants. The structural work presented here in combination with biochemical experiments using artificial tRNA or artificial lipid substrates reveals the tRNA acceptor stem, the aminoacyl moiety, and the polar head group of PG as the main determinants for substrate recognition. A mutagenesis approach yielded the complementary amino acid determinants of tRNA interaction. These results have broad implications for the design of L-PGS and A-PGS inhibitors that could render microbial pathogens more susceptible to antimicrobial compounds.
en
Structures of two bacterial resistance factors mediating tRNA-dependent aminoacylation of phosphatidylglycerol with lysine or alanine.
Article
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URL
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oai:repository.helmholtz-hzi.de:10033/6210492019-08-30T11:35:39Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Jäger, Jens
author
Marwitz, Sebastian
author
Tiefenau, Jana
author
Rasch, Janine
author
Shevchuk, Olga
author
Kugler, Christian
author
Goldmann, Torsten
author
Steinert, Michael
department
Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
2017-08-08T11:25:42Z
2014-01
Human lung tissue explants reveal novel interactions during Legionella pneumophila infections. 2014, 82 (1):275-85 Infect. Immun.
1098-5522
24166955
10.1128/IAI.00703-13
http://hdl.handle.net/10033/621049
Infection and immunity
Histological and clinical investigations describe late stages of Legionnaires' disease but cannot characterize early events of human infection. Cellular or rodent infection models lack the complexity of tissue or have nonhuman backgrounds. Therefore, we developed and applied a novel model for Legionella pneumophila infection comprising living human lung tissue. We stimulated lung explants with L. pneumophila strains and outer membrane vesicles (OMVs) to analyze tissue damage, bacterial replication, and localization as well as the transcriptional response of infected tissue. Interestingly, we found that extracellular adhesion of L. pneumophila to the entire alveolar lining precedes bacterial invasion and replication in recruited macrophages. In contrast, OMVs predominantly bound to alveolar macrophages. Specific damage to septa and epithelia increased over 48 h and was stronger in wild-type-infected and OMV-treated samples than in samples infected with the replication-deficient, type IVB secretion-deficient DotA(-) strain. Transcriptome analysis of lung tissue explants revealed a differential regulation of 2,499 genes after infection. The transcriptional response included the upregulation of uteroglobin and the downregulation of the macrophage receptor with collagenous structure (MARCO). Immunohistochemistry confirmed the downregulation of MARCO at sites of pathogen-induced tissue destruction. Neither host factor has ever been described in the context of L. pneumophila infections. This work demonstrates that the tissue explant model reproduces realistic features of Legionnaires' disease and reveals new functions for bacterial OMVs during infection. Our model allows us to characterize early steps of human infection which otherwise are not feasible for investigations.
en
Human lung tissue explants reveal novel interactions during Legionella pneumophila infections.
Article
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oai:repository.helmholtz-hzi.de:10033/6211142019-08-30T11:37:24Zcom_10033_6832col_10033_6833
Helmholtz Zentrum für Infektionsforschung Repository
author
Schniederjans, Monika
author
Koska, Michal
author
Häussler, Susanne
department
Helmholtz Centre for infection researchGmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
2017-09-22T08:16:17Z
2017-09-05
Transcriptional and mutational profiling of an aminoglycoside resistant Pseudomonas aeruginosa small colony variant. 2017 Antimicrob. Agents Chemother.
1098-6596
28874369
10.1128/AAC.01178-17
http://hdl.handle.net/10033/621114
Antimicrobial agents and chemotherapy
Pseudomonas aeruginosa is a major causative agent of both acute and chronic infections. Although aminoglycoside antibiotics are very potent drugs to fight such infections, antibiotic failure is steadily increasing mainly due to increasing resistance of the bacteria. Many molecular mechanisms that determine resistance such as acquisition of genes encoding for aminoglycoside-inactivating enzymes or overexpression of efflux pumps have been elucidated. However, there are additional, less-well described mechanisms of aminoglycoside resistance. In this study we have profiled a clinical tobramycin resistant P. aeruginosa strain that exhibited a small colony variant (SCV) phenotype. Both, the resistance and the colony morphology phenotypes were lost upon passaging the isolate under rich medium conditions. Transcriptional and mutational profiling revealed that the SCV harbored activating mutations in the two two-component systems AmgRS and PmrAB. Introduction of these mutations singularly into the type strain PA14 conferred tobramycin and colistin resistance, respectively. However, their combined introduction had an additive effect on the tobramycin resistance phenotype. Activation of the AmgRS system slightly reduced the colony size of the PA14 wild-type, whereas the simultaneous overexpression of gacA, the response regulator of the GacSA two component system, further reduced colony size. In conclusion, we uncovered combinatorial influences of two-component systems on clinically relevant phenotypes, such as resistance and the expression of the SCV phenotype. Our results clearly demonstrate that combined activation of P. aeruginosa two-component systems exhibit pleiotropic effects with unforeseen consequences.
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Transcriptional and mutational profiling of an aminoglycoside resistant Pseudomonas aeruginosa small colony variant.
Article
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https://hzi.openrepository.com/bitstream/10033/621114/1/Schniederjans%20et%20al.pdf
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