Enhanced extracellular Bacillus stearothermophilus alpha-amylase production in Bacillus subtilis by balancing the entire secretion process in an optimal strain

摘要

A novel strategy that combines host strain optimization with balancing of the entire secretion process has been used to enhance the extracellular production of Bacillus stearothermophilus a-amylase (AmySA) in Bacillus subtilis. B. subtilis strain WS9, which lacks six extracellular proteases, was selected as the most suitable host for recombinant AmySA expression. This strain was further modified by inactivating the hrcA gene, which increases the expression of intracellular molecular chaperones. Then, with co-expressing the transport channel component SecYEG complex, the entire secretion process of a-amylase was balanced through the signal peptide SPRpmG. The resulting recombinant strain (WHS9GSAB) efficiently produced AmySA, showing an extracellular AmySA activity of 2835.1 U/mL during shake-flask cultivation. This was 2.9-fold greater than that of the original strain WS11YSA (976.9 U/mL). Cultivation of WHS9GSAB in a 3-L fermenter produced an extracellular AmySA activity of 35779.5 U/mL at 93 h, with a high productivity of 384.7 U/mL.h. This represents the highest level of re-combinant alpha-amylase production in B. subtilis reported to date. The expression strategy developed in this study may be useful for both the industrial production of alpha-amylases and improving the extracellular production of other recombinant proteins in B. subtilis.