Efficient secretory expression of Bacillus stearothermophilus α/β-cyclodextrin glycosyltransferase in Bacillus subtilis.

Lingqia Su, Yunfei Li, Jing Wu

Journal of biotechnology 2021 Apr 10

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Bacillus stearothermophilus α/β-cyclodextrin glycosyltransferase (α/β-CGTase) is an excellent transglycosylase with broad potential for food application, but its expression level is low in Bacillus subtilis. In this study, the optimal signal peptide for α/β-CGTase expression was screened from 173 signal peptides in B. subtilis WS11. The α/β-CGTase activity in a 3-L fermentor reached 151.93 U⋅ mL-1, but substantial amounts of inclusion bodies were produced. The N-terminal 12 amino acids of α/β-CGTase were then replaced with the N-terminal 15 amino acids of a β-CGTase from the same family that has a high percentage of disorder-promoting amino acids. As a result, the inclusion bodies were significantly reduced, and the enzyme activity increased to 249.35 U mL-1, 2.3 times that of the strain constructed previously. Finally, the ppsE and sfp genes of B. subtilis WS11, which are related to lipopeptide biosurfactant synthesis, were knocked out to produce B. subtilis WS13. When B. subtilis WS13 was used to produce α/β-CGTase in a 3-L fermentor, 70 % less defoaming agent was required than with B. subtilis WS11. Furthermore, enzyme production and growth of WS13 were equivalent to those of WS11. This study is of great significance for future research to efficiently scale-up production of α/β-CGTase. Copyright © 2021 Elsevier B.V. All rights reserved.

Citation

Lingqia Su, Yunfei Li, Jing Wu. Efficient secretory expression of Bacillus stearothermophilus α/β-cyclodextrin glycosyltransferase in Bacillus subtilis. Journal of biotechnology. 2021 Apr 10;331:74-82