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KMID : 1234420100380020151
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Korean Journal of Microbiololgy and Biotechnology
2010 Volume.38 No. 2 p.151 ~ p.157
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Enzymatic Properties of Barley α-Amylase Chimeric Enzymes Produced by Staggered Extension Process
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Kim Tae-Jip
Choi Seung-Ho
Jang Myoung-Uoon
Park Jung-Mi
Birte Svensson
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Abstract
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Barley malt produces two different α-amylase isozymes (AMY1 and AMY2), which share up to 80% of amino acid sequence identity with each other. However, their enzymatic properties differ remarkably. In this study, five chimeric enzymes between AMY1 and 2 were constructed by staggered extension process (StEP) technique, and their enzymatic properties were characterized. According to the results, chimeric AMY-D2, D8, and E12 showed the mixed or intermediate types of calcium-dependent activity between AMY1 and 2. Meanwhile, only AMY-E10 chimera could be significantly inhibited by barley α-amylase/subtilisin inhibitor (BASI) protein. Chimera AMY-C6 showed the same calcium-dependency as AMY1, while AMY-E10 was closely similar to AMY2. As a result, it can be proposed that some amino acid residues in the region II, III, and IV of barley α-amylases can play very important roles in the interaction with BASI, and those in III, V, VI, and VII may partly affect on the calcium-dependent activity.
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KEYWORD
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Barley α-amylase isozymes, staggered extension process(StEP), chimeric enzymes, calciumdependent activity, Pichia pastoris
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