A gene encoding for an alpha-amylase from thermophilic Bacillus sp. strain TS-23 and its expression in Escherichia coli |
| |
Authors: | Lin L L Hsu W H Chu W S |
| |
Institution: | Department of Food Nutrition, Hung Kuang Institute of Medical and Nursing Technology, Shalu, Taichung, Taiwan. |
| |
Abstract: | An alpha-amylase gene from Bacillus sp. strain TS-23 was cloned and expressed by using its own promoter on the recombinant plasmid pTS917 in Escherichia coli. A cell fractionation experiment revealed that approximately 60% of the amylase activity was in the periplasmic space. Analysis and activity staining of the concentrated supernatant fraction by SDS-polyacrylamide gel electrophoresis showed an apparent protein band with a mol. wt of approximately 65,000. The amylase gene (amyA) consisted of an open reading frame of 1,845 bp encoding a protein of 613 amino acids with a calculated mol. wt of 69,543. The predicted amino acid sequence showed high homology with Bacillus species, E. coli and Salmonella typhimurium alpha-amylases. Deletion of 96 amino acids from the C-terminal portion of the amylase did not result in the loss of amylolytic activity. The truncated amylase, deletion of the first 50 amino acids from the N-terminus, was overexpressed in E. coli system and refolded to yield an activable enzyme. |
| |
Keywords: | |
本文献已被 PubMed 等数据库收录! |
|