14 research outputs found
Use of a Sec signal peptide library from Bacillus subtilis for the optimization of cutinase secretion in Corynebacterium glutamicum
The twin-arginine translocation (Tat) pathway: Investigation of substrate-translocase interactions during Tat-dependent protein translocation in E. coli.
Entwicklung und Anwendung von Biosensoren fĂŒr die Optimierung der Sec-abhĂ€ngigen heterologen Proteinsekretion in Corynebacterium glutamicum
Optimization of Sec-dependent heterologous protein secretion in Corynebacterium glutamicum
Combinatorial impact of Sec signal peptides from Bacillus subtilis and bioprocess conditions on heterologous cutinase secretion by Corynebacterium glutamicum
The impact of Sec signal peptides (SPs) from Bacillus subtilis in combination with isopropylâÎČâ dâ1âthiogalactopyranoside concentration and feeding profile was investigated for heterologous protein secretion performance by Corynebacterium glutamicum using cutinase as model enzyme. Based on a comprehensive data set of about 150 benchâscale bioreactor cultivations in fedâbatch mode and choosing the cutinase yield as objective, it was shown that relative secretion performance for bioprocesses remains very similar, irrespective of the applied SP enabling Secâmediated cutinase secretion. However, to achieve the maximal absolute cutinase yield, careful adjustment of bioprocess conditions was found to be necessary. A modelâbased, twoâstep multiple regression approach resembled the collected data in a comprehensive way. The corresponding results suggest that the choice of the heterologous Sec SP and its interaction with the adjusted exponential feeding profile is highly relevant to maximize absolute cutinase yield in this study. For example, the impact of Sec SP is high at low growth rates and low at high growth rates. However, promising Sec SPs could be inferred from less complex batch cultivations. The extensive data were also evaluated in terms of cutinase productivity, highlighting the wellâknown tradeâoff between yield and productivity in bioprocess development in detail. Conclusively, only the right combination of target protein, Sec SP, and bioprocess conditions is the key to success
Proper selection of signal peptides for heterologous expression and secretion: Challenging, but worthwile
Use of a Sec signal peptide library from Bacillus subtilis for the optimization of cutinase secretion in Corynebacterium glutamicum
Technical bulk enzymes represent a huge market, and the extracellular production of such enzymes is favorable due to lowered cost for product recovery. Protein secretion can be achieved via general secretion (Sec) pathway. Specific sequences, signal peptides (SPs), are necessary to direct the target protein into the translocation machinery. For example, >150 Sec-specific SPs have been identified for Bacillus subtilis alone. As the best SP for a target protein of choice cannot be predicted a priori, screening of homologous SPs has been shown to be a powerful tool for different expression organisms. While SP libraries between closely related species were successfully applied to optimize recombinant protein secretion, this was not investigated for distantly related species. Therefore, in this study a Sec SP library from low-GC firmicutes B. subtilis is investigated to optimize protein secretion in high-GC actinobacterium Corynebacterium glutamicum using cutinase from Fusarium solani pisi as model protein. [...