Characterization of Chimeric Enzymes Constructed between Two Distinct α-Amylase cDNAs from Cultured Rice Cells

Abstract

  Cultured cells of rice (Oryza sativa cv Sasanishiki) produce two α-amylase isozymes, AMY-I and AMY-III. Using a bacterial expression system, eight chimeric genes constructed with various combination of AMY-I and AMY-III cDNA fragments were expressed, and each recombinant chimeric protein was characterized. Four of the eight recombinant enzymes having region c (one of the four regions having unconserved base sequences between AMY-I and AMY-III cDNAs) of AMY-I showed the same enzyme characteristics as that of native AMY-I, which had high temperature optimum at 50°C. The other four chimeric proteins carrying region c of AMY-III showed the AMY-III type characteristics, which were a low temperature optimum at 25°C and susceptibility to a higher maltooligosaccharide (G17) substrate. The unconserved region c is involved in the decision of the characteristic of AMY-I or AMY-III.