Screening, identification, purification and homologous modeling of marine cold-active alpha-amylase

Cold-active α-amylase is essential in industrial production. However, the number of cold-active α-amylases available for use is limited. Screening microbial strains would lay the groundwork for the future development of the food and pharmaceutical industries.

To screen microbial strains for cold-active α-amylase based on physiological and biochemical identification, as well as homology modelling.

Cold-active α-amylase strains were screened from water and mud obtained from the Yellow Sea. Colony morphology, Gram staining, scanning electron microscopy and transmission electron microscopy, physiological and biochemical identification and 16S rRNA gene analysis were used to identify strains. A series of steps, including DEAE-anion exchange column chromatography, SephadexG-100 column chromatography, and SDS-PAGE electrophoresis, were used to produce cold-active α-amylase of relatively high purity. Finally, the homology of amylase was modeled to explore the structure and activity site of the enzyme.

The named dsh19-1 strain of cold-active α-amylase was screened and identified as Bacillus. The cold-active α-amylase produced by Bacillus was named AmyD-1. The protein with PDB sequence number 5A2B was found to have 40.6% homology with AmyD-1. The verification score of the 3-D model was 137.07 points. We discovered that the six sites are potential sites for amylase to decompose starch by building a 3-D AmyD-1 model. AmyD-1 has a molecular weight of 1515 bp, and hydrogen bonding may be the primary interaction force between AmyD-1 and glucose molecules.

A cold-active α-amylase produced by Bacillus strain dsh19-1 was successfully obtained and named AmyD-1. This enzyme has potential uses in the food and pharmaceutical industries.