Optimization of the Thermal Activity and Stability of Hyperthermophilic α-amylase ApkA

doi:10.13560/j.cnki.biotech.bull.1985.2017-0172

Abstract

Abstract: This work aims to obtain α-amylase with improved thermal activity and stability. Based on the structure analysis of hyperthermophilic α-amylase ApkA and the most thermo-stable α-amylase PFA，a Zn²⁺-binding site mutant ApkAdsK152H/A166C was constructed by introducing the Zn²⁺-binding site of PFA into ApkA. The mutant ApkAdsK152H/A166C exhibited effective increase in terms of thermal activity and stability. The optimal temperature of the mutant increased from 90℃ to 100℃ and the corresponding specific activity was 5 201.08 U/mg. The half-life of ApkAdsK152H/A166C prolonged from 5 h to 10 h while incubated at 90℃，and from 7.5 min to 80 min at 100℃. The results of Zn²⁺ content measurement in recombinant α-amylases confirmed that ApkAdsK152H/A166C bound with one Zn²⁺ ion. These results suggested that the introduction of Zn²⁺-binding site improved the thermal activity and stability of ApkA.

Key words: hyperthermophilic α-amylase, Zn²⁺-binding site, site-directed mutagenesis, thermo-activity, thermostability

ZENG Jing, GUO Jian-jun, YUAN Lin, YANG Gang, CHEN Jun. Optimization of the Thermal Activity and Stability of Hyperthermophilic α-amylase ApkA[J]. Biotechnology Bulletin, 2017, 33(8): 192-198.

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