2024-03-29T11:28:59Zhttp://repository.helmholtz-hzi.de/oai/requestoai:repository.helmholtz-hzi.de:10033/1399622019-08-30T11:33:57Zcom_10033_620533col_10033_620538
The intra- and extracellular proteome of Aspergillus niger growing on defined medium with xylose or maltose as carbon substrate.
Lu, Xin
Sun, Jibin
Nimtz, Manfred
Wissing, Josef
Zeng, An-Ping
Rinas, Ursula
Helmholtz Center for Infection Research, Inhoffenstr, Braunschweig, Germany.
The filamentous fungus Aspergillus niger is well-known as a producer of primary metabolites and extracellular proteins. For example, glucoamylase is the most efficiently secreted protein of Aspergillus niger, thus the homologous glucoamylase (glaA) promoter as well as the glaA signal sequence are widely used for heterologous protein production. Xylose is known to strongly repress glaA expression while maltose is a potent inducer of glaA promoter controlled genes. For a more profound understanding of A. niger physiology, a comprehensive analysis of the intra- and extracellular proteome of Aspergillus niger AB1.13 growing on defined medium with xylose or maltose as carbon substrate was carried out using 2-D gel electrophoresis/Maldi-ToF and nano-HPLC MS/MS.
2011-08-17T13:57:00Z
2011-08-17T13:57:00Z
2010
Article
The intra- and extracellular proteome of Aspergillus niger growing on defined medium with xylose or maltose as carbon substrate. 2010, 9:23 Microb. Cell Fact.
1475-2859
20406453
10.1186/1475-2859-9-23
http://hdl.handle.net/10033/139962
Microbial cell factories
en
oai:repository.helmholtz-hzi.de:10033/1903392019-08-30T11:33:28Zcom_10033_620533col_10033_620538
Improving the prediction of Pseudomonas putida mt-2 growth kinetics with the use of a gene expression regulation model of the TOL plasmid
Koutinas, Michalis
Kiparissides, Alexandros
Lam, Ming-Chi
Silva-Rocha, Rafael
Godinho, Miguel
de Lorenzo, Victor
Martins dos Santos, Vitor A.P.
Pistikopoulos, Efstratios N.
Mantalaris, Athanasios
Helmholtz Center for Infection Research (HZI), 38124 Braunschweig, Germany.
2011-11-22T15:39:44Z
2011-11-22T15:39:44Z
2011-11-22T15:39:44Z
Article
Improving the prediction of Pseudomonas putida mt-2 growth kinetics with the use of a gene expression regulation model of the TOL plasmid 2011, 55 (2):108 Biochemical Engineering Journal
1369703X
10.1016/j.bej.2011.03.012
http://hdl.handle.net/10033/190339
Biochemical Engineering Journal
null
http://linkinghub.elsevier.com/retrieve/pii/S1369703X11000829
oai:repository.helmholtz-hzi.de:10033/2162972019-08-30T11:33:57Zcom_10033_620533col_10033_620538
A viability assay for Candida albicans based on the electron transfer mediator 2,6-dichlorophenolindophenol.
Hassan, Rabeay Y A
Bilitewski, Ursula
Biological Systems Analysis Group, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany.
Candida albicans is an opportunistic fungal pathogen with comparably high respiratory activity. Thus, we established a viability test based on 2,6-dichlorophenolindophenol (DCIP), a membrane-permeable electron transfer agent. NADH dehydrogenases catalyze the reduction of DCIP by NADH, and the enzymatic activity can be determined either electrochemically via oxidation reactions of DCIP or photometrically. Among the specific respiratory chain inhibitors, only the complex I inhibitor rotenone decreased the DCIP signal from C. albicans, leaving residual activity of approximately 30%. Thus, the DCIP-reducing activity of C. albicans was largely dependent on complex I activity. C. albicans is closely related to the complex I-negative yeast Saccharomyces cerevisiae, which had previously been used in DCIP viability assays. Via comparative studies, in which we included the pathogenic complex I-negative yeast Candida glabrata, we could define assay conditions that allow a distinction of complex I-negative and -positive organisms. Basal levels of DCIP turnover by S.cerevisiae and C. glabrata were only 30% of those obtained from C. albicans but could be increased to the C. albicans level by adding glucose. No significant increases were observed with galactose. DCIP reduction rates from C. albicans were not further increased by any carbon source.
2012-03-22T14:02:04Z
2012-03-22T14:02:04Z
2011-12-01
Article
A viability assay for Candida albicans based on the electron transfer mediator 2,6-dichlorophenolindophenol. 2011, 419 (1):26-32 Anal. Biochem.
1096-0309
21864496
10.1016/j.ab.2011.07.025
http://hdl.handle.net/10033/216297
Analytical biochemistry
en
Archived with thanks to Analytical biochemistry
oai:repository.helmholtz-hzi.de:10033/2697162019-08-30T11:32:41Zcom_10033_620533col_10033_620538
The fungicide fludioxonil antagonizes fluconazole activity in the human fungal pathogen Candida albicans.
Buschart, Anna
Burakowska, Anna
Bilitewski, Ursula
Biological Systems Analysis, Helmholtz Centre for Infection Research, Braunschweig, Germany.
The fungicide fludioxonil is widely used in agriculture. Residua of this fungicide are occasionally detected in fruits and can therefore be ingested by humans. The human fungal pathogen Candida albicans expresses the target of fludioxonil, Nik1p, a type III histidine kinase involved in stress response. Inhibition of yeast and hyphae growth was hardly observable after treatment of C. albicans SC5314 with fludioxonil. As a side effect, however, we observed a concentration-dependent induction of the expression of the genes CDR1 and CDR2, encoding ATP-binding cassette (ABC) transporters. This was independent of the presence of the target of fludioxonil as induction was also observed in a Δnik1 deletion mutant. Deletion of the CDR1 gene aggravated the inhibition of germ tube formation by fludioxonil, indicating that, in the wild-type, the fungicide was discharged from the cell by Cdr1p. Cdr1p is also known as a resistance factor of C. albicans against the commonly used antimycotic fluconazole. Thus, the effect of concurrent exposure to fludioxonil and known cargoes of ABC transporters on their extrusion and the growth of C. albicans was examined. Pre-incubation with fludioxonil decreased the export rate of rhodamine 6G. The resistance to fluconazole was increased by fludioxonil, independently of Nik1p. Therefore, exposure of C. albicans to fludioxonil may lead to increased resistance to fluconazole treatment.
2013-02-18T15:19:02Z
2013-02-18T15:19:02Z
2012-12
Article
The fungicide fludioxonil antagonizes fluconazole activity in the human fungal pathogen Candida albicans. 2012, 61 (Pt 12):1696-703 J. Med. Microbiol.
1473-5644
22918865
10.1099/jmm.0.050963-0
http://hdl.handle.net/10033/269716
Journal of medical microbiology
en
Archived with thanks to Journal of medical microbiology
oai:repository.helmholtz-hzi.de:10033/2837732019-08-30T11:36:05Zcom_10033_620533col_10033_620538
Involvement of the mitogen activated protein kinase Hog1p in the response of Candida albicans to iron availability.
Kaba, Hani Ej
Nimtz, Manfred
Müller, Peter P
Bilitewski, Ursula
ABSTRACT: BACKGROUND: Iron is an essential nutrient for almost all organisms, and generating iron limiting conditions for pathogens is one of the host defense strategies against microbial infections. Excess of iron can be toxic; therefore, iron uptake is tightly controlled. The high affinity iron uptake system of the opportunistic pathogenic yeast Candida albicans has been shown to be essential for virulence. Several transcription factors and regulators of iron uptake genes were identified, but the knowledge of signaling pathways is still limited. Gene expression profiling of the Deltahog1 deletion mutant indicated an involvement of the mitogen activated protein (MAP) kinase Hog1p. However, the function of Hog1p in the response of C. albicans to iron availability was not studied in detail. Thus, we analyzed phenotypic and molecular responses of C. albicans to different iron concentrations particularly with respect to the activity of the Hog1p MAP kinase module. RESULTS: We observed flocculation of yeast cells, when the iron ion concentration was equal to or higher than 5 muM. This phenotype was dependent on the MAP kinase Hog1p and the corresponding MAP kinase kinase Pbs2p. Moreover, high extracellular iron ion concentrations led to hyper-phosphorylation of Hog1p. We determined lower amounts of multicopper ferroxidase (MCFO) proteins and lower ferric reductase activity, when the iron ion concentration in the medium was increased. This effect was also observed for the Deltahog1 mutant. However, the amounts of MCFO proteins and the cell surface ferric reductase activity were increased in the Deltahog1 in comparison to wild type cells. This effect was independent of iron availability in growth media. CONCLUSIONS: In C. albicans, the MAP kinase Hog1p is part of the network regulating the response of the organism to iron availability. Hog1p was transiently phosphorylated under high iron concentrations and was essential for a flocculent phenotype. Furthermore, deletion of HOG1 led to increased levels of components of the reductive iron uptake system in comparison to the wild-type, independent of iron concentrations in the media. However, the additional induction of this system by low iron concentrations was independent of HOG1.
2013-04-24T12:06:10Z
2013-04-24T12:06:10Z
2013-01-24
Article
Involvement of the mitogen activated protein kinase Hog1p in the response of Candida albicans to iron availability. 2013, 13 (1):16 BMC Microbiol.
1471-2180
23347662
10.1186/1471-2180-13-16
http://hdl.handle.net/10033/283773
BMC microbiology
Archived with thanks to BMC microbiology
oai:repository.helmholtz-hzi.de:10033/2954842019-08-30T11:33:57Zcom_10033_620533col_10033_620538
Direct electrochemical determination of Candida albicans activity.
Hassan, Rabeay Y A
Bilitewski, Ursula
Helmholtz Centre for Infection Research, Working Group BiSA, Inhoffenstr. 7, 38124 Braunschweig, Germany; Microanalysis Lab, Applied Organic Chemistry Department, National Research Centre (NRC), Eltahrir Street, 12311-Dokki, Cairo, Egypt. Electronic address: rabeayy@yhaoo.com.
Despite advances made in the field, rapid detection methods for the human pathogen Candida albicans are still missing. In this regard, bio-electrochemical systems including electrochemical sensors and biosensors satisfy the increasing demand for rapid, reliable, and direct microbial analyses. In this study, the bioelectrochemical characteristics of C. albicans were investigated for use in an analytical system that determines the viability of the organisms. The electrochemical responses of viable and non-viable cells of C. albicans and Saccharomyces cerevisiae were monitored. Cyclic voltammograms (CV) showed an irreversible oxidation peak at about 750mV that accounts for viable cells. The peak current increased at viable cell numbers ranging from 3×10(5) to 1.6×10(7)cells/ml, indicating that the amount of viable cells can be accurately quantified. To elucidate the underlying electron transfer processes, the influence of electron transfer chain (ETC) - inhibitors on the electrochemical behavior of the two organisms were investigated. Inhibition of the function of classical respiratory chain (CRC) led to a decrease in the electrochemical response, whereas the oxidation current increased when the alternative oxidase (AOX) pathway was blocked by salicylhydroxamic acid (SHA). Blocking the AOX pathway improved the electrochemical performance, suggesting an involvement in the CRC, with cytochrome c oxidase (COX) as a relevant protein complex. Mutants, in which components of COX were deleted, showed a lower electro-activity than the wild-type strain. Particularly, deletion of subunit COX5a almost completely abolished the electrochemical signal. We believe that this work can be utilized for the development of early detection assays and opens the door for new technological developments in the field of C. albicans.
2013-07-08T13:36:57Z
2013-07-08T13:36:57Z
2013-05-15
Article
Direct electrochemical determination of Candida albicans activity. 2013, 49C:192-198 Biosens Bioelectron
1873-4235
23747360
10.1016/j.bios.2013.05.015
http://hdl.handle.net/10033/295484
Biosensors & bioelectronics
Archived with thanks to Biosensors & bioelectronics
oai:repository.helmholtz-hzi.de:10033/2974772019-08-30T11:34:22Zcom_10033_620533col_10033_620538
Identification of inhibitors of yeast-to-hyphae transition in Candida albicans by a reporter screening assay.
Heintz-Buschart, Anna
Eickhoff, Holger
Hohn, Erwin
Bilitewski, Ursula
Department of Biological Systems Analysis, Helmholtz Centre for Infection Research, Inhoffenstr. 7, 38124 Braunschweig, Germany.
Candida albicans is one of the most common opportunistic fungal pathogens, causing life-threatening disease in immunocompromised patients. As it is not primarily a pathogen, but can exist in a commensal state, we aimed at the identification of new anti-infective compounds which do not eradicate the fungus, but primarily disable a virulence determinant. The yeast–hyphae-dimorphism of C. albicans is considered a major contributor to fungal disease, as mutants locked into either yeast or hyphal state have been shown to be less virulent in the mouse-model. We devised a high-throughput screening procedure which allows us to find inhibitors of the induction of hyphae. Hyphae-formation was induced by nitrogen starvation at 37 °C and neutral pH in a reporter strain, which couples promoter activity of the hyphae-specific HWP1 to β-galactosidase expression. In a pilot screening of 720 novel synthetic compounds, we identified substances which inhibited the outgrowth of germ tubes. They belonged to chemical classes not yet known for antimycotic properties, namely methyl aryl-oxazoline carboxylates, dihydrobenzo[d]isoxazolones and thiazolo[4,5-e]benzoisoxazoles. In conclusion we developed a novel screening assay, which addresses the morphological switch from the yeast form of C. albicans to its hyphal form and identified novel chemical structures with activity against C. albicans.
2013-08-07T14:04:32Z
2013-08-07T14:04:32Z
2013-03-10
Article
Identification of inhibitors of yeast-to-hyphae transition in Candida albicans by a reporter screening assay. 2013, 164 (1):137-42 J. Biotechnol.
1873-4863
23262131
10.1016/j.jbiotec.2012.12.004
http://hdl.handle.net/10033/297477
Journal of biotechnology
en
Archived with thanks to Journal of biotechnology
oai:repository.helmholtz-hzi.de:10033/3050742019-08-30T11:30:32Zcom_10033_620533col_10033_620538
Deletion of the HAMP domains from the histidine kinase CaNik1p of Candida albicans or treatment with fungicides activates the MAP kinase Hog1p in S. cerevisiae transformants.
El-Mowafy, Mohammed
Bahgat, Mahmoud M
Bilitewski, Ursula
Microorganisms use two-component signal transduction (TCST) systems to regulate the response of the organism to changes of environmental conditions. Such systems are absent from mammalian cells and are thus of interest as drug targets. Fungal TCST systems are usually composed of a hybrid histidine kinase, comprising the histidine kinase (HisKA) domain and a receiver domain, a histidine phosphotransfer protein and a response regulator. Among the 11 groups of fungal histidine kinases, group III histidine kinases are of particular relevance as they are essential for the activity of different groups of fungicides. A characteristic feature is the N-terminal amino acid repeat domain comprising multiple HAMP domains, of which the function is still largely unknown. In Candida albicans, a fungal human pathogen, three histidine kinases were identified, of which CaNik1p is a group III histidine kinase. Heterologous expression of this protein in Sacchromyces cerevisiae conferred susceptibility to different fungicides. Fungicide activity was associated with phosphorylation of the mitogen activated protein kinase Hog1p.
2013-11-07T12:05:13Z
2013-11-07T12:05:13Z
2013-09-17
Article
Deletion of the HAMP domains from the histidine kinase CaNik1p of Candida albicans or treatment with fungicides activates the MAP kinase Hog1p in S. cerevisiae transformants. 2013, 13 (1):209 BMC Microbiol.
1471-2180
24044701
10.1186/1471-2180-13-209
http://hdl.handle.net/10033/305074
BMC microbiology
Archived with thanks to BMC microbiology
oai:repository.helmholtz-hzi.de:10033/6210542018-06-13T02:46:39Zcom_10033_620533col_10033_620538
Virus-like particle production with yeast: ultrastructural and immunocytochemical insights into Pichia pastoris producing high levels of the Hepatitis B surface antigen
Lünsdorf, Heinrich
Gurramkonda, Chandrasekhar
Adnan, Ahmad
Khanna, Navin
Rinas, Ursula
Abstract Background A protective immune response against Hepatitis B infection can be obtained through the administration of a single viral polypeptide, the Hepatitis B surface antigen (HBsAg). Thus, the Hepatitis B vaccine is generated through the utilization of recombinant DNA technology, preferentially by using yeast-based expression systems. However, the polypeptide needs to assemble into spherical particles, so-called virus-like particles (VLPs), to elicit the required protective immune response. So far, no clear evidence has been presented showing whether HBsAg assembles in vivo inside the yeast cell into VLPs or later in vitro during down-stream processing and purification. Results High level production of HBsAg was carried out with recombinant Pichia pastoris using the methanol inducible AOX1 expression system. The recombinant vaccine was isolated in form of VLPs after several down-stream steps from detergent-treated cell lysates. Search for the intracellular localization of the antigen using electron microscopic studies in combination with immunogold labeling revealed the presence of HBsAg in an extended endoplasmic reticulum where it was found to assemble into defined multi-layered, lamellar structures. The distance between two layers was determined as ~6 nm indicating that these lamellas represent monolayers of well-ordered HBsAg subunits. We did not find any evidence for the presence of VLPs within the endoplasmic reticulum or other parts of the yeast cell. Conclusions It is concluded that high level production and intrinsic slow HBsAg VLP assembly kinetics are leading to retention and accumulation of the antigen in the endoplasmic reticulum where it assembles at least partly into defined lamellar structures. Further transport of HBsAg to the Golgi apparatus is impaired thus leading to secretory pathway disfunction and the formation of an extended endoplasmic reticulum which bulges into irregular cloud-shaped formations. As VLPs were not found within the cells it is concluded that the VLP assembly process must take place during down-stream processing after detergent-mediated disassembly of HBsAg lamellas and subsequent reassembly of HBsAg into spherical VLPs.
2017-08-14T10:15:19Z
2017-08-14T10:15:19Z
2011-06-26
2015-09-04T08:22:20Z
Journal Article
Microbial Cell Factories. 2011 Jun 26;10(1):48
http://dx.doi.org/10.1186/1475-2859-10-48
http://hdl.handle.net/10033/621054
en
Lünsdorf et al; licensee BioMed Central Ltd.
oai:repository.helmholtz-hzi.de:10033/6207842019-08-30T11:26:12Zcom_10033_620533col_10033_620538
Deletion of the HAMP domains from the histidine kinase CaNik1p of Candida albicans or treatment with fungicides activates the MAP kinase Hog1p in S. cerevisiae transformants
El-Mowafy, Mohammed
Bahgat, Mahmoud M
Bilitewski, Ursula
Abstract Background Microorganisms use two-component signal transduction (TCST) systems to regulate the response of the organism to changes of environmental conditions. Such systems are absent from mammalian cells and are thus of interest as drug targets. Fungal TCST systems are usually composed of a hybrid histidine kinase, comprising the histidine kinase (HisKA) domain and a receiver domain, a histidine phosphotransfer protein and a response regulator. Among the 11 groups of fungal histidine kinases, group III histidine kinases are of particular relevance as they are essential for the activity of different groups of fungicides. A characteristic feature is the N-terminal amino acid repeat domain comprising multiple HAMP domains, of which the function is still largely unknown. In Candida albicans, a fungal human pathogen, three histidine kinases were identified, of which CaNik1p is a group III histidine kinase. Heterologous expression of this protein in Sacchromyces cerevisiae conferred susceptibility to different fungicides. Fungicide activity was associated with phosphorylation of the mitogen activated protein kinase Hog1p. Results We have constructed mutated versions of CaNik1p, from which either all HAMP domains were deleted (CaNik1pΔHAMP) or in which the histidine kinase or the receiver domains were not-functional. Expression of CaNIK1ΔHAMP in S. cerevisiae led to severe growth inhibition. Normal growth could be restored by either replacing the phosphate-accepting histidine residue in CaNik1pΔHAMP or by expressing CaNIK1ΔHAMP in S. cerevisiae mutants, in which single genes encoding several components of the HOG pathway were deleted. Expression of proteins with non-functional histidine kinase or receiver domains resulted in complete loss of susceptibility to antifungals, such as fludioxonil. Conditions leading to growth inhibition of transformants also led to phosphorylation of the MAP kinase Hog1p. Conclusion Our results show that functional histidine kinase and receiver domains of CaNik1p were essential for antifungal susceptibility and for activation of the Hog1p. Moreover, for the first time we show that deletion of all HAMP domains from CaNik1p led to activation of Hog1p without an external stimulus. This phenotype was similar to the effects obtained upon treatment with fungicides, as in both cases growth inhibition correlated with Hog1p activation and was dependent on the functionality of the conserved phosphate-accepting histidine residue.
2017-01-27T11:38:20Z
2017-01-27T11:38:20Z
2013-09-17
2015-09-04T08:26:59Z
Journal Article
BMC Microbiology. 2013 Sep 17;13(1):209
http://dx.doi.org/10.1186/1471-2180-13-209
http://hdl.handle.net/10033/620784
en
El-Mowafy et al.; licensee BioMed Central Ltd.
oai:repository.helmholtz-hzi.de:10033/5992592019-11-21T13:17:44Zcom_10033_620533col_10033_620538col_10033_620538
Regulation of Candida albicans Interaction with Macrophages through the Activation of HOG Pathway by Genistein
Cui, Shuna
Hassan, Rabeay
Heintz-Buschart, Anna
Bilitewski, Ursula
Helmholtz Centre for infection research, Inhoffenstr. 7, D-38124 Braunschweig, Germany.
2016-02-25T15:46:22Z
2016-02-25T15:46:22Z
2016-01-28
Article
Regulation of Candida albicans Interaction with Macrophages through the Activation of HOG Pathway by Genistein 2016, 21 (2):162 Molecules
1420-3049
26828477
10.3390/molecules21020162
http://hdl.handle.net/10033/599259
Molecules
http://www.mdpi.com/1420-3049/21/2/162
oai:repository.helmholtz-hzi.de:10033/6214692019-08-30T11:32:15Zcom_10033_620533col_10033_620538
Self-assembly and biological activities of ionic liquid crystals derived from aromatic amino acids.
Neidhardt, Manuel M
Schmitt, Katharina
BARO, ANGELIKA
Schneider, Carmen
Bilitewski, Ursula
LASCHAT, SABINE
Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
The self-assembly of amino acid-derived ionic liquid crystals (ILCs) into lamellar or micellar-like aggregates suggests that they might interact with biological membranes. To get some insight, guanidinium chlorides derived from the natural l-amino acids phenylalanine (Phe), tyrosine (Tyr) and 3,4-dihydroxyphenylalanine (DOPA) were synthesized and their mesomorphic properties were investigated via polarizing optical microscopy (POM), differential scanning calorimetry (DSC) and X-ray diffraction (SAXS, WAXS). Mesophase types depended on the number of alkoxy side chains. Phe- and Tyr-based ILCs with one and two side chains, respectively, self-assembled into smectic A bilayers (SmA2), while Dopa-derived ILCs with three side chains formed columnar (Colh) mesophases. The mesophase ranges for Phe ILCs increased steadily with side chain length, for Tyr- and Dopa-based ILCs, however, size matching effects were observed. To clarify whether the mesomorphic behaviour has an impact on biological properties, cytotoxic and antibacterial activities of the ILCs were studied. Phe and Tyr ILCs exhibited much higher cytotoxicities (against the L-929 mouse fibroblast cell line) and/or antibacterial activities (against Staphylococcus aureus) than Dopa ILCs, which were mostly inactive. Furthermore, within each series, the side chain length largely influenced the biological activity. Thus, the bulk mesophase behaviour appeared to correlate with the biological properties, in particular, the interactions with membranes, as shown by measuring the intracellular Ca2+ concentration in human monocytic U937 cells after treatment with the amino acid-based ILCs.
2018-09-05T11:34:44Z
2018-09-05T11:34:44Z
2018-08-08
Article
1463-9084
30043016
10.1039/c8cp03404d
http://hdl.handle.net/10033/621469
http://creativecommons.org/licenses/by-nc-sa/3.0/us/
Attribution-NonCommercial-ShareAlike 3.0 United States
Physical chemistry chemical physics : PCCP
oai:repository.helmholtz-hzi.de:10033/6215282019-08-30T11:29:41Zcom_10033_620533col_10033_620538
Triazole-based cross-linkers in radical polymerization processes: tuning mechanical properties of poly(acrylamide) and poly( -dimethylacrylamide) hydrogels
Götz, Tobias
Schädel, Nicole
Petri, Nadja
Kirchhof, Manuel
Bilitewski, Ursula
Tovar, Günter E. M.
Laschat, Sabine
Southan, Alexander
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
Triazole-based cross-linkers with different spacer lengths and different functional end groups (acrylamides,
methacrylamides, maleimides and vinylsulfonamides) were synthesized, investigated for cytotoxic and
antibacterial activity, and incorporated into poly(acrylamide) (PAAm) and poly(N,N-dimethylacrylamide)
(PDMAAm) hydrogels by free-radical polymerization. Hydrogels prepared with different cross-linkers and
cross-linker contents between 0.2% and 1.0% were compared by gel yields, equilibrium degrees of swelling
(S) and storage moduli (G0). Generally with increasing cross-linker content, G0 values of the hydrogels
increased, while S values decreased. The different polymerizable cross-linker end groups resulted in
a decrease of G0 in the following order for cross-linkers with C4 spacers: acrylamide > maleimide >
methacrylamide > vinylsulfonamide. Longer cross-linker alkyl spacer lengths caused an increase in G0 and
a decrease in S. Independent of the cross-linker used, a universal correlation between G0 and equilibrium
polymer volume fraction f was found. For PAAm hydrogels, G0 ranged between 4 kPa and 23 kPa and f
between 0.07 and 0.14. For PDMAAm hydrogels, G0 ranged between 0.1 kPa and 4.9 kPa and f between
0.02 and 0.06. The collected data were used to establish an empirical model to predict G0 depending on f.
G0 of PAAm and PDMAAm hydrogels is given by G0 ¼ 4034 kPa f2.66 and G0 ¼ 4297 kPa f2.46, respectively.
2018-11-01T13:21:22Z
2018-11-01T13:21:22Z
Article
2046-2069
10.1039/C8RA07145D
http://hdl.handle.net/10033/621528
http://xlink.rsc.org/?DOI=C8RA07145D
http://creativecommons.org/licenses/by-nc-sa/3.0/us/
Attribution-NonCommercial-ShareAlike 3.0 United States
8
60
34743
34753
RSC Advances
oai:repository.helmholtz-hzi.de:10033/6220262019-11-23T01:59:46Zcom_10033_620533col_10033_620538
Regulation of Candida albicans Interaction with Macrophages through the Activation of HOG Pathway by Genistein
Cui, Shuna
Hassan, Rabeay
Heintz-Buschart, Anna
Bilitewski, Ursula
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
he severity of infections caused by Candida albicans, the most common opportunistic human fungal pathogen, needs rapid and effective antifungal treatments. One of the effective ways is to control the virulence factors of the pathogen. Therefore, the current study examined the effects of genistein, a natural isoflavone present in soybeans, on C. albicans. The genistein-treated C. albicans cells were then exposed to macrophages. Although no inhibition effect on the growth rates of C. albicans was noted an enhancement of the immune response to macrophages has been observed, indicated by phagocytosis and release of cytokines TNF-α and IL-10. The effect of genistein on the enhanced phagocytosis can be mimicked by the fungicides fludioxonil or iprodione, which inhibit the histidine kinase Cos1p and lead to activation of HOG pathway. The western blot results showed a clear phosphorylation of Hog1p in the wild type strain of C. albicans after incubation with genistein. In addition, effects of genistein on the phosphorylation of Hog1p in the histidine kinase mutants Δcos1 and Δsln1 were also observed. Our results thus indicate a new bio-activity of genistein on C. albicans by activation of the HOG pathway of the human pathogen C. albicans.
2019-11-22T14:47:18Z
2019-11-22T14:47:18Z
2016-01-28
Article
Molecules. 2016 Jan 28;21(2):162. doi: 10.3390/molecules21020162.
1420-3049
26828477
10.3390/molecules21020162
http://hdl.handle.net/10033/622026
Molecules
en
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
MDPI AG
21
2
162
oai:repository.helmholtz-hzi.de:10033/6222052020-03-14T01:59:39Zcom_10033_620533col_10033_620538
A NanoLuc luciferase-based assay enabling the real-time analysis of protein secretion and injection by bacterial type III secretion systems.
Westerhausen, Sibel
Nowak, Melanie
Torres-Vargas, Claudia E
Bilitewski, Ursula
Bohn, Erwin
Grin, Iwan
Wagner, Samuel
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
Salmonella
Luciferase
Type III secretion system
effector proteins
protein injection
protein secretion
The elucidation of the molecular mechanisms of secretion through bacterial protein secretion systems is impeded by a shortage of assays to quantitatively assess secretion kinetics. Also the analysis of the biological role of these secretion systems as well as the identification of inhibitors targeting these systems would greatly benefit from the availability of a simple, quick and quantitative assay to monitor principle secretion and injection into host cells. Here, we present a versatile solution to this need, utilizing the small and very bright NanoLuc luciferase to assess the function of the type III secretion system encoded by Salmonella pathogenicity island 1. Type III secretion substrate-NanoLuc fusions are readily secreted into the culture supernatant, where they can be quantified by luminometry after removal of bacteria. The NanoLuc-based secretion assay features a very high signal-to-noise ratio and sensitivity down to the nanolitre scale. The assay enables monitoring of secretion kinetics and is adaptable to a high throughput screening format in 384-well microplates. We further developed a split NanoLuc-based assay that enables the real-time monitoring of type III secretion-dependent injection of effector-HiBiT fusions into host cells stably expressing the complementing NanoLuc-LgBiT.
2020-03-13T10:23:53Z
2020-03-13T10:23:53Z
2020-02-18
Article
Mol Microbiol. 2020 Feb 18. doi: 10.1111/mmi.14490.
32068313
10.1111/mmi.14490
http://hdl.handle.net/10033/622205
1365-2958
Molecular microbiology
en
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
Wiley
Molecular microbiology
England
oai:repository.helmholtz-hzi.de:10033/6222612020-05-14T01:36:37Zcom_10033_620533col_10033_620538
Inhibition of Respiration of Candida albicans by Small Molecules Increases Phagocytosis Efficacy by Macrophages.
Cui, Shuna
Li, Minghui
Hassan, Rabeay Y A
Heintz-Buschart, Anna
Wang, Junsong
Bilitewski, Ursula
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
antimycin A
complex III
metabolism
phagocytosis
respiratory chain
Candida albicans adapts to various conditions in different body niches by regulating gene expression, protein synthesis, and metabolic pathways. These adaptive reactions not only allow survival but also influence the interaction with host cells, which is governed by the composition and structure of the fungal cell wall. Numerous studies had shown linkages between mitochondrial functionality, cell wall integrity and structure, and pathogenicity. Thus, we decided to inhibit single complexes of the respiratory chain of C. albicans and to analyze the resultant interaction with macrophages via their phagocytic activity. Remarkably, inhibition of the fungal bc1 complex by antimycin A increased phagocytosis, which correlated with an increased accessibility of β-glucans. To contribute to mechanistic insights, we performed metabolic studies, which highlighted significant changes in the abundance of constituents of the plasma membrane. Collectively, our results reinforce the strong linkage between fungal energy metabolism and other components of fungal physiology, which also determine the vulnerability to immune defense reactions.IMPORTANCE The yeast Candida albicans is one of the major fungal human pathogens, for which new therapeutic approaches are required. We aimed at enhancements of the phagocytosis efficacy of macrophages by targeting the cell wall structure of C. albicans, as the coverage of the β-glucan layer by mannans is one of the immune escape mechanisms of the fungus. We unambiguously show that inhibition of the fungal bc1 complex correlates with increased accessibilities of β-glucans and improved phagocytosis efficiency. Metabolic studies proved not only the known direct effects on reactive oxygen species (ROS) production and fermentative pathways but also the clear downregulation of the ergosterol pathway and upregulation of unsaturated fatty acids. The changed composition of the plasma membrane could also influence the interaction with the overlying cell wall. Thus, our work highlights the far-reaching relevance of energy metabolism, indirectly also for host-pathogen interactions, without affecting viability.
2020-05-13T13:35:25Z
2020-05-13T13:35:25Z
2020-04-15
Article
mSphere. 2020 Apr 15;5(2). pii: 5/2/e00016-20. doi: 10.1128/mSphere.00016-20.
32295866
10.1128/mSphere.00016-20
http://hdl.handle.net/10033/622261
2379-5042
mSphere
en
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
ASM
5
2
mSphere
United States
oai:repository.helmholtz-hzi.de:10033/6222602020-05-14T01:36:30Zcom_10033_620533col_10033_620538
Inhibition of Respiration of Candida albicans by Small Molecules Increases Phagocytosis Efficacy by Macrophages.
Cui, Shuna
Li, Minghui
Hassan, Rabeay Y A
Heintz-Buschart, Anna
Wang, Junsong
Bilitewski, Ursula
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstraße 7, 38124 Braunschweig, Germany.
antimycin A
complex III
metabolism
phagocytosis
respiratory chain
Candida albicans adapts to various conditions in different body niches by regulating gene expression, protein synthesis, and metabolic pathways. These adaptive reactions not only allow survival but also influence the interaction with host cells, which is governed by the composition and structure of the fungal cell wall. Numerous studies had shown linkages between mitochondrial functionality, cell wall integrity and structure, and pathogenicity. Thus, we decided to inhibit single complexes of the respiratory chain of C. albicans and to analyze the resultant interaction with macrophages via their phagocytic activity. Remarkably, inhibition of the fungal bc1 complex by antimycin A increased phagocytosis, which correlated with an increased accessibility of β-glucans. To contribute to mechanistic insights, we performed metabolic studies, which highlighted significant changes in the abundance of constituents of the plasma membrane. Collectively, our results reinforce the strong linkage between fungal energy metabolism and other components of fungal physiology, which also determine the vulnerability to immune defense reactions.IMPORTANCE The yeast Candida albicans is one of the major fungal human pathogens, for which new therapeutic approaches are required. We aimed at enhancements of the phagocytosis efficacy of macrophages by targeting the cell wall structure of C. albicans, as the coverage of the β-glucan layer by mannans is one of the immune escape mechanisms of the fungus. We unambiguously show that inhibition of the fungal bc1 complex correlates with increased accessibilities of β-glucans and improved phagocytosis efficiency. Metabolic studies proved not only the known direct effects on reactive oxygen species (ROS) production and fermentative pathways but also the clear downregulation of the ergosterol pathway and upregulation of unsaturated fatty acids. The changed composition of the plasma membrane could also influence the interaction with the overlying cell wall. Thus, our work highlights the far-reaching relevance of energy metabolism, indirectly also for host-pathogen interactions, without affecting viability.
2020-05-13T13:32:11Z
2020-05-13T13:32:11Z
2020-04-15
Article
mSphere. 2020 Apr 15;5(2). pii: 5/2/e00016-20. doi: 10.1128/mSphere.00016-20.
32295866
10.1128/mSphere.00016-20
http://hdl.handle.net/10033/622260
2379-5042
mSphere
en
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
American Society of Microbiology
5
2
mSphere
United States
oai:repository.helmholtz-hzi.de:10033/6223772020-08-04T02:35:32Zcom_10033_620533col_10033_620538
Polyhalonitrobutadienes as Versatile Building Blocks for the Biotargeted Synthesis of Substituted N-Heterocyclic Compounds.
Zapol'skii, Viktor A
Bilitewski, Ursula
Kupiec, Sören R
Ramming, Isabell
Kaufmann, Dieter E
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
heterocyclization
medicinal chemistry
nitrogen heterocycles
nucleophilic substitution
polyhalonitrobutadienes
Substituted nitrogen heterocycles are structural key units in many important pharmaceuticals. A new synthetic approach towards heterocyclic compounds displaying antibacterial activity against Staphylococcus aureus or cytotoxic activity has been developed. The selective synthesis of a series of 64 new N-heterocycles from the three nitrobutadienes 2-nitroperchloro-1,3-butadiene, 4-bromotetrachloro-2-nitro-1,3-butadiene and (Z)-1,1,4-trichloro-2,4-dinitrobuta-1,3-diene proved feasible. Their reactions with N-, O- and S-nucleophiles provide rapid access to push-pull substituted benzoxazolines, benzimidazolines, imidazolidines, thiazolidinones, pyrazoles, pyrimidines, pyridopyrimidines, benzoquinolines, isothiazoles, dihydroisoxazoles, and thiophenes with unique substitution patterns. Antibacterial activities of 64 synthesized compounds were examined. Additionally, seven compounds (thiazolidinone, nitropyrimidine, indole, pyridopyrimidine, and thiophene derivatives) exhibited a significant cytotoxicity with IC50-values from 1.05 to 20.1 µM. In conclusion, it was demonstrated that polyhalonitrobutadienes have an interesting potential as structural backbones for a variety of highly functionalized, pharmaceutically active heterocycles.
2020-08-03T13:16:56Z
2020-08-03T13:16:56Z
2020-06-21
Article
Molecules. 2020;25(12):2863. Published 2020 Jun 21. doi:10.3390/molecules25122863.
32575902
10.3390/molecules25122863
http://hdl.handle.net/10033/622377
1420-3049
Molecules (Basel, Switzerland)
en
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
MDPI
25
12
Molecules (Basel, Switzerland)
Switzerland
oai:repository.helmholtz-hzi.de:10033/6224912020-10-01T03:02:55Zcom_10033_620533col_10033_620538
Synthesis and Biological Evaluation of a Library of AGE-Related Amino Acid Triazole Crosslinkers
Icik, Esra
Jolly, Anthony
Löffler, Paul
Agelidis, Nektarios
Bugdayci, Bakiye
Altevogt, Luca
Bilitewski, Ursula
Baro, Angelika
LASCHAT, SABINE
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
Physical and Theoretical Chemistry
Organic Chemistry
Three N‐Boc‐protected amino acids, l‐serine, l‐aspartic, and l‐glutamic acid, were either converted into their methyl azidoalkanoates or various alkynes via Bestmann‐Ohira strategy or via reaction with propargylamine and propargyl bromide, respectively. The Cu‐catalyzed click reaction provided a library of amino acid based triazoles, which were further N‐methylated to triazolium iodides or deprotected and precipitated as free amino acid triazole dihydrochlorides. The biological properties of all derivatives were investigated by cytotoxicity assay (against L929 mouse fibroblasts) and broth microdilution method (E. coli ΔTolC and S. aureus). First results reveal complete inactivity for triazolium iodides with cell viabilities and microbial growths nearly 100 %, indicating them as possible analogs of advanced glycation endproducts (AGEs).
2020-09-30T12:59:45Z
2020-09-30T12:59:45Z
2020-08-10
Article
Eur J Org Chem 2020;2020(33):5368-5379 doi:10.1002/ejoc.202000811.
1434-193X
10.1002/ejoc.202000811
http://hdl.handle.net/10033/622491
European Journal of Organic Chemistry
en
http://doi.wiley.com/10.1002/tdm_license_1.1
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
Wiley-VCH
European Journal of Organic Chemistry
oai:repository.helmholtz-hzi.de:10033/6226032020-11-24T01:42:17Zcom_10033_620533col_10033_620538
Buchwald-Hartwig versus Microwave-Assisted Amination of Chloroquinolines: En Route to the Pyoverdin Chromophore
Seubert, Philipp
Freund, Marcel
Rudolf, Richard
Lin, Yulin
Altevogt, Luca
Bilitewski, Ursula
Baro, Angelika
Laschat, Sabine
HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
Amination
aminonitroquinolines
Buchwald-Hartwig amination
microwave heating
pyoverdins
siderophores
The reaction of 2-chloro-3-nitroquinoline and a series of amines and aminoalkanoates under
basic microwave-mediated conditions and Buchwald-Hartwig amination conditions is
reported. The microwave irradiation favored the reaction with amines, resulting in yields up to
80%, while amino acid functionalization gave yields comparable to those of BuchwaldHartwig amination. (2R)-4-[(6,7-dimethoxy-3-nitroquinolinyl)amino]-2-hydroxybutanoate
could be successfully cyclized to the pyoverdin chromophore, a subunit of siderophores.
2020-11-23T08:38:04Z
2020-11-23T08:38:04Z
2020-07-22
Article
Synlett 2020 31 12 doi: 10.1055/s-0040-1707810.
09365214
10.1055/s-0040-1707810
http://hdl.handle.net/10033/622603
14372096
Synlett
2-s2.0-85087912934
SCOPUS_ID:85087912934
en
http://creativecommons.org/licenses/by-nc-sa/4.0/
Attribution-NonCommercial-ShareAlike 4.0 International
Thieme Verlag
31
12
1177
1181
Synlett