Abstract
In this study, first, β-mannanase gene man derived from Bacillus amyloliquefaciens CGMCC1.857 was cloned and expressed in Bacillus subtilis 168 to generate B. subtilis M1. However, the extracellular β-mannanase activity of B. subtilis M1 was not very high. To further increase extracellular β-mannanase extracytoplasmic molecular chaperone, PrsA lipoprotein was tandem expressed with man gene in B. subtilis 168 to yield B. subtilis M2. The secretion of β-mannanase of B. subtilis M2 was enhanced by 15.4%, compared with the control B. subtilis M1. Subsequently, process optimization strategies were also developed to enhance β-mannanase production by B. subtilis 168 M2. It was noted that the optimal temperature for β-mannanase production (25°C) was different from the optimal growth temperature (37°C) for B. subtilis. Based on these findings, a two-stage temperature control strategy was proposed where the bacterial culture was maintained at 37°C for the first 12 h to obtain a high rate of cell growth, followed by lowering the temperature to 25°C to enhance β-mannanase production. Using this strategy, the extracellular β-mannanase activity reached 5016 ± 167 U/ml at about 36 h, which was 19.1% greater than the best result obtained using a constant temperature (25°C). The result of this study showed that PrsA lipoprotein overexpression and two-stage temperature control strategy were more efficient for β-mannanase fermentation in B. subtilis.
Published Version
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