Cloning and extracellular expression of a raw starch digesting α-amylase (Blamy-I) and its application in bioethanol production from a non-conventional source of starch
메타 데이터
바이오화학분류
바이오플라스틱
플라스틱
바이오정밀화학
용매
화학제품
연료
화장품용 기능성소재
계면활성제⁄증점제
의료용 화학소재
식품첨가제
논문
Cloning and extracellular expression of a raw starch digesting α-amylase (Blamy-I) and its application in bioethanol production from a non-conventional source of starch
<P>The aim of this study was to clone and efficiently express a raw starch‐digesting α‐amylase enzyme in the culture media and also to investigate the potential application of this recombinant enzyme in the digestion of non‐conventional raw starch for bioethanol production. A raw starch digesting α‐amylase gene isolated from <I>Bacillus licheniformis</I> strain AS08E was cloned and extracellularly expressed in <I>E. coli</I> cells using the native signal peptide. The mature recombinant α‐amylase (Blamy‐I) consisting of 483 amino acid residues was found to be homogenous with a mass of 55.3 kDa (by SDS‐PAGE analysis) and a predicted pI of 6.05. Structural and functional analysis of Blamy‐I revealed the presence of an extra Ca<SUP>2+</SUP>‐binding region between the A and C domains responsible for higher thermostability of this enzyme. The statistical optimization of <I>E. coli</I> culture conditions resulted in an approximately eightfold increase in extracellular expression of Blamy‐I as compared to its production under non‐optimized conditions. Blamy‐I demonstrated optimum enzyme activity at 80 °C and pH 10.0, and efficiently hydrolyzed raw starch isolated from a non‐conventional, underutilized jack fruit seeds. Further utilization of this starch for bioethanol production using Blamy‐I and <I>Saccharomyces cerevisiae</I> also proved to be highly promising.</P>