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KMID : 0545120120220081077
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Journal of Microbiology and Biotechnology
2012 Volume.22 No. 8 p.1077 ~ p.1083
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Investigations on Possible Roles of C-Terminal Propeptide of a Ca-Independent α-Amylase from Bacillus
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Ali Salimi
Fatemeh Yousefi
Marzieh Ghollasi
Sara Daneshjou
Hesam Tavoli
Sirous Ghobadi
Khosro Khajeh
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Abstract
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Previously, an extracellular α-amylase (BKA) had been purified from the culture of Bacillus sp. KR8104. Subsequently, the crystal structure of the active enzyme revealed a 422 amino acids polypeptide. In this study, the bka was cloned into E. coli, which encoded a polypeptide of 659 amino acids including two additional fragments: one 44 residues N-terminal fragment and another 193 residues C-terminal fragment. In order to investigate the role of the C-terminal fragment, two constructs with and without this region [BKAΔ(N44) and BKAΔ(N44C193)] were designed and expressed in E. coli BL21. The optimum pH, thermal stability, and the end-products of starch hydrolysis were found to be similar in both constructs. The Km and Vmax values for BKAΔ(N44) were lower than BKAΔ(N44C193), using either starch or ethylidene-blocked 4-nitrophenylmaltoheptaoside as a substrate.
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KEYWORD
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α-Amylase, Bacillus sp. KR-8104, Biochemical characterization, Secretion, Truncation
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