Enhanced extracellular α-amylase production in Brevibacillus choshinensis by optimizing extracellular degradation and folding environment

• Published in: Journal of industrial microbiology & biotechnology Vol. 49; no. 1
• Main Authors: Yao, Dongbang, Zhang, Kang, Zhu, Xuyang, Su, Lingqia, Wu, Jing
• Format: Journal Article
• Language: English
• Published: Germany Oxford University Press 20.01.2022
• Subjects: alpha-Amylases - biosynthesis; alpha-Amylases - genetics; Bioreactors; Brevibacillus - genetics; Brevibacillus - metabolism; Fermentation, Cell Culture and Bioengineering; Protein Folding; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Brevibacillus choshinensis; α-Amylase; Protease; 3-l Fermenter fermentation; Chaperone
• ISSN: 1367-5435; 1476-5535
• DOI: 10.1093/jimb/kuab061

A strategy for optimizing the extracellular degradation and folding environment of Brevibacillus choshinensis has been used to enhance the extracellular production of recombinant α-amylase. First, a gene (bcp) encoding an extracellular protease and another encoding an extracellular chaperone (prsC) were identified in the genome of B. choshinensis HPD31-SP3. Then, the effect of extracellular protein degradation on recombinant α-amylase production was investigated by establishing a CRISPR/Cas9n system to knock out bcp. The effect of extracellular folding capacity was investigated separately by coexpressing extracellular chaperones genes from different sources (prsA, prsC, prsL, prsQ) in B. choshinensis. The final recombinant strain (BCPPSQ), which coexpressed prsQ in a genetic background lacking bcp, produced an extracellular α-amylase activity of 6940.9 U/ml during shake-flask cultivation. This was 2.1-fold greater than that of the original strain BCWPS (3367.9 U/ml). Cultivation of BCPPSQ in a 3-l fermenter produced an extracellular α-amylase activity of 17925.6 U/ml at 72 h, which was 7.6-fold greater than that of BCWPS (2358.1 U/ml). This strategy demonstrates its great potential in enhancing extracellular α-amylase production in B. choshinensis. What's more, this study provides a strategic reference for improving the extracellular production of other recombinant proteins in B. choshinensis.