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Construction of a highly active secretory expression system via an engineered dual promoter and a highly efficient signal peptide in Bacillus subtilis

Paper ID Volume ID Publish Year Pages File Format Full-Text
32925 44951 2016 8 PDF Available
Title
Construction of a highly active secretory expression system via an engineered dual promoter and a highly efficient signal peptide in Bacillus subtilis
Abstract

•PgsiB–PHpaII was prominent among the single and dual promoters evaluated.•SPYncM was selected from 19 signal peptides to optimize the secretion efficiency.•The capacity of the expression system was demonstrated with batch fermentation.

A strong promoter and highly efficient signal peptides are essential for the secretory overproduction of recombinant proteins in Bacillus subtilis. To enhance the limited overexpression capability of natural promoters, various strategies for promoter engineering have been developed and used to construct gene expression systems in B. subtilis and other hosts. By applying a semi-rational approach for promoter engineering, a series of expression plasmids containing single and dual promoters were constructed using aminopeptidase (AP) with an intrinsic signal peptide as the reporter protein. Of the single and dual promoters investigated, the dual promoter PgsiB–PHpaII gave the best performance. To optimize secretion efficiency, the signal peptide YncM was selected after screening a library containing 19 different Sec-type signal peptides. The AP activity detected in the supernatants of a recombinant strain containing the plasmid pBSG24-YncM was as high as 88.86 U/mL. The capacity of the expression plasmid pBSG24-YncM was also evaluated with batch fermentation in a 5-L fermentor. Increased production of AP (205 U/mL, equal to 1.7 g/L) was achieved after 45 h of fermentation. These results suggest that this expression system can be used for high-level protein expression in B. subtilis.

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Construction of a highly active secretory expression system via an engineered dual promoter and a highly efficient signal peptide in Bacillus subtilis
Publisher
Database: Elsevier - ScienceDirect
Journal: New Biotechnology - Volume 33, Issue 3, 25 May 2016, Pages 372–379
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering