北京棒杆菌天冬氨酸激酶突变体P184Q的酶学性质表征

Characterization of Enzymatic Properties of Mutant P184Q of Aspartate Kinase from Corynebacterium pekinense

通过分析谷氨酸棒杆菌天冬氨酸激酶(AK)的结构,筛选可能影响别构抑制剂结合的Pro184位点,对其进行饱和定点突变,成功筛选出突变菌株P184Q.酶学性质研究表明:突变体P184Q的V_(max)比野生型(WT)提高了3倍;n=1.39,低于WT的值(2.6),正协同性下降,同时Km值减小,对底物的亲和力增大;P184Q最适pH=6.5,最适反应温度为25℃,半衰期为2.8h;P184Q对金属离子和有机溶剂均表现出良好的抗性,解除抑制剂苏氨酸和甲硫氨酸、赖氨酸和甲硫氨酸、苏氨酸和赖氨酸、赖氨酸对酶活力的抑制作用.

By analyzing the structure of aspartate kinase（AK）,we screened the Pro184 site that might affect the binding of allosteric inhibitors,and the mutant strain P184Q was successfully screened by saturation site-directed mutagenesis.The enzymatic properties show that the V_（max） of P184Q was 3 times higher than that of wide-type（WT）,n=1.39 indicating that the positive cooperativity of the substrate decreased,the Kmvalue decreased simultaneously,and the affinity of the substrate increased.The optimum pH of P184Q was 6.5.The optimum temperature of P184Q was 25 ℃.The half-life period was 2.8 h.Futhermore,P184Q how strong resistance to metal ions,organic solvent and inhibitors Thr＋Met,Lys＋Met,Thr＋Lys and Lys.