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Published on: August 13, 2011
Expression, biochemical and structural characterization of high-specific-activity β-amylase from Bacillus aryabhattai
Xuguo Duan1, Qiuyu Zhu2, Xinyi Zhang2
1College of Light Industry and Food Engineering, Nanjing Forestry University, Nanjing, 210037, Jiangsu, China. xgduan@njfu.edu.cn.
Background:
β-amylase (EC 3.2.1.2) is an exo-enzyme that shows high specificity for cleaving the α-1,4-glucosidic linkage of starch from the non-reducing end, thereby liberating maltose. In this study, we heterologously expressed and characterized a novel β-amylase from Bacillus aryabhattai.
Results:
The amino acid-sequence alignment showed that the enzyme shared the highest sequence identity with β-amylase from Bacillus flexus (80.73%) followed by Bacillus cereus (71.38%). Structural comparison revealed the existence of an additional starch-binding domain (SBD) at the C-terminus of B. aryabhattai β-amylase, which is notably different from plant β-amylases. The recombinant enzyme purified 4.7-fold to homogeneity, with a molecular weight of ~ 57.6 kDa and maximal activity at pH 6.5 and 50 °C. Notably, the enzyme exhibited the highest specific activity (3798.9 U/mg) among reported mesothermal microbial β-amylases and the highest specificity for soluble starch, followed by corn starch. Kinetic analysis showed that the Km and kcat values were 9.9 mg/mL and 116961.1 s- 1, respectively. The optimal reaction conditions to produce maltose from starch resulted in a maximal yield of 87.0%. Moreover, molecular docking suggested that B. aryabhattai β-amylase could efficiently recognize and hydrolyze maltotetraose substrate.
Conclusions:
These results suggested that B. aryabhattai β-amylase could be a potential candidate for use in the industrial production of maltose from starch.
