Comparative proteomic analysis of high cell density cultivations with two recombinant Bacillus megaterium strains for the production of a heterologous dextransucrase.
Average rating
Cast your vote
You can rate an item by clicking the amount of stars they wish to award to this item.
When enough users have cast their vote on this item, the average rating will also be shown.
Star rating
Your vote was cast
Thank you for your feedback
Thank you for your feedback
Issue Date
2006
Metadata
Show full item recordAbstract
High cell density cultivations were performed under identical conditions for two Bacillus megaterium strains (MS941 and WH320), both carrying a heterologous dextransucrase (dsrS) gene under the control of the xylA promoter. At characteristic points of the cultivations (end of batch, initial feeding, before and after induction) the proteome was analyzed based on two dimensional gel electrophoresis and mass spectrometric protein identification using the protein database "bmegMEC.v2" recently made available.High expression but no secretion of DsrS was found for the chemical mutant WH320 whereas for MS 941, a defined protease deficient mutant of the same parent strain (DSM319), not even expression of DsrS could be detected. The proteomic analysis resulted in the identification of proteins involved in different cellular pathways such as in central carbon and overflow metabolism, in protein synthesis, protein secretion and degradation, in cell wall metabolism, in cell division and sporulation, in membrane transport and in stress responses.The two strains exhibited considerable variations in expression levels of specific proteins during the different phases of the cultivation process, whereas induction of DsrS production had, in general, little effect. The largely differing behaviour of the two strains with regard to DsrS expression can be attributed, at least in part, to changes observed in the proteome which predominantly concern biosynthetic enzymes and proteins belonging to the membrane translocation system, which were strongly down-regulated at high cell densities in MS941 compared with WH320. At the same time a cell envelope-associated quality control protease and two peptidoglycan-binding proteins related to cell wall turnover were strongly expressed in MS941 but not found in WH320. However, to further explain the very different physiological responses of the two strains to the same cultivation conditions, it is necessary to identify the mutated genes in WH320 in addition to the known lacZ.In view of the results of this proteomic study it seems that at high cell density conditions and hence low growth rates MS941, in contrast to WH320, does not maintain a vegetative growth which is essential for the expression of the foreign dsrS gene by using the xylA promoter. It is conceivable that applications of a promoter which is highly active under nutrient-limited cultivation conditions is necessary, at least for MS941, for the overexpression of recombinant genes in such B. megaterium fed-batch cultivation process. However to obtain a heterologous protein in secreted and properly folded form stills remains a big challenge.Citation
Proteome Sci 2006, 4:19PubMed ID
17022804Type
ArticleLanguage
enISSN
1477-5956ae974a485f413a2113503eed53cd6c53
10.1186/1477-5956-4-19
Scopus Count
Related articles
- Proteome analysis of a recombinant Bacillus megaterium strain during heterologous production of a glucosyltransferase.
- Authors: Wang W, Hollmann R, Fürch T, Nimtz M, Malten M, Jahn D, Deckwer WD
- Issue date: 2005 May 31
- Production and secretion of recombinant Leuconostoc mesenteroides dextransucrase DsrS in Bacillus megaterium.
- Authors: Malten M, Hollmann R, Deckwer WD, Jahn D
- Issue date: 2005 Jan 20
- Coexpression of the type I signal peptidase gene sipM increases recombinant protein production and export in Bacillus megaterium MS941.
- Authors: Malten M, Nahrstedt H, Meinhardt F, Jahn D
- Issue date: 2005 Sep 5
- Codon optimized Thermobifida fusca hydrolase secreted by Bacillus megaterium.
- Authors: Yang Y, Malten M, Grote A, Jahn D, Deckwer WD
- Issue date: 2007 Mar 1
- Recombinant production of the antibody fragment D1.3 scFv with different Bacillus strains.
- Authors: Lakowitz A, Krull R, Biedendieck R
- Issue date: 2017 Jan 23