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Construction and Enzymatic Properties of Double Mutant M372I-T379S of Aspartokinase from Corynebacterium pekinense#br#
GAO Yunna, HAN Caijing, ZHAN Dongling, FANG Li, MIN Weihong
Journal of Jilin University Science Edition. 2019, 57 (5):
1267-1274.
In order to construct mutants with high enzyme activity of aspartokinase (AK) and weaken or relieve the feedback inhibition of Lys (lysine), mutants M372I,T379S and M372I-T379S were constructed by site\|directed mutagenesis and high-throughput screening techniques. The wild type (WT) and mutants were induced, expressed and characterized by enzymatic properties. The results show that compared with the WTAK, the Vmax of mutants M372I,T379S and M372I-T379S AK increases 13.77,15.02 and 15.60 times, respectively, and the values of Km and n are lower than that of WTAK, the optimum pH values of mutants M372I,T379S and M372I-T379S AK are increased slightly, which are 8.0,8.5,8.5, and the half\|life period of mutants are prolonged by 1.0,0.9 and 2.3 h, respectively. The optimal temperature of M372I-T379S AK is 30 ℃, which is higher than WT AK 2 ℃. When the concentration is 1—10 mmol/L, the mutants weaken or partially relieve the feedback inhibition of Lys inhibitor.
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