Investigations on possible roles of C-terminal propeptide of a Ca-independent α-amylase from bacillus.
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 V(max) values for BKAΔ(N44) were lower than BKAΔ(N44C193), using either starch or ethylidene-blocked 4-nitrophenylmaltoheptaoside as a substrate.
Department of Biochemistry, Faculty of Biological Science, Tarbiat Modares University, Tehran, Iran.
Ali
A
Salimi
Fatemeh
F
Yousefi
Marzieh
M
Ghollasi
Sara
S
Daneshjou
Hesam
H
Tavoli
Sirous
S
Ghobadi
Khosro
K
Khajeh
D000681:Amylases / Q000235:genetics / Q000378:metabolism*; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013213:Starch / Q000378:metabolism; D013696:Temperature
D011993:Recombinant Fusion Proteins; D013213:Starch; D000681:Amylases