IMPROVED THERMOSTABILITY OF A BACILLUS ALPHA-AMYLASE BY DELETION OF AN ARGININE-GLYCINE RESIDUE IS CAUSED BY ENHANCED CALCIUM-BINDING

alpha-Amylase from alkaliphilic Bacillus KSM-1378 (LAMY) is a novel se mi-alkaline enzyme which has a high specific activity, a value 5-fold higher than that of a Bacillus licheniformis enzyme at alkaline pH. Th ermostability of this enzyme could be improved by deletion of the Arg1 81-Gly182 residue by means of site-directed mutagenesis. The wild-type and engineered LAMYs were very similar with respect to specific activ ity, pH-activity curve, temperature-activity curve, susceptibility to inhibitors, and pattern of hydrolysis products from soluble starch and maltooligosaccharides. However, the engineered enzyme also acquired i ncreased pH stability and resistance to sodium dodecyl sulfate and esp ecially chelating reagents, such as ethylenediaminetetraacetate and et hyleneglycol-bis (beta-aminoethylether)tetraacetate. This is the first report that thermostability of alpha-amylase is improved by enhanced calcium binding to the enzyme molecule, (C) 1998 Academic Press.