Enhanced extracellular expression of α-Amylase DL3-4-1 in Bacillus subtilis via systematic screening of optimal signal peptides
2021
Abstract Amylases have been widely applied for starch saccharification in the food, paper, pharmaceutical, and textile industries. In the present study, α-amylase DL-3-4−1 from B. amyloliquefaciens was successfully expressed in B. subtilis. The enzymatic activity exhibited high stability in a pH range from 4.0 to 11.0 and moderate thermostability. Furthermore, excellent stability was observed when incubated with metal ions, detergents, inhibitors, and organic solvents. Resistance against ethanol, urea, and guanidine−HCl was also observed. More importantly, a strategy to enhance the extracellular expression of α-amylase DL-3-4−1 was developed using 173 Sec-type signal peptides (SPs) as a library based on a fast and sequence-independent method. Fourteen SPs were identified with significantly increased yields of DL-3-4−1 via a high-throughput assay. Specifically, the enzymatic activity of the α-amylase-producing strain containing the signal peptide SP5D10 (YomL) was 3.74-fold higher than that of the strain with the wild type signal peptide (87.30 U/OD). In brief, the maximum specific activity of an α-amylase-producing strain containing the YomL signal peptide was 326.45 U/OD. This is the first report of an α-amylase that exhibits ethanol, urea, and guanidine−HCl tolerance. Hence, this α-amylase represents a promising candidate for future use in industry.
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