Val22对恶臭假单胞菌扁桃酸消旋酶催化活性的影响  

The effect of Val22 on the catalytic activity of mandelate racemase from Pseudomonas putida

作　　者：梅卓航[1] 刘击[1] 于洪巍[1] 

机构地区：[1]浙江大学生物工程研究所,浙江杭州310027

出　　处：《微生物学通报》2011年第7期982-987,共6页Microbiology China

摘　　要：恶臭假单胞菌扁桃酸消旋酶的Val22位于20s环状结构上,是与底物结合相关的氨基酸之一。其中Val被替换为Arg后酶活性下降了75.9%。除了酶与底物疏水作用减弱以外,静电排斥作用增强也可能引起活性的下降。利用分子动力学模拟对酶与底物的米氏复合物进行分析,结果表明:突变后第22位氨基酸侧链与底物的静电势从0.036 kJ/mol升高至0.124 kJ/mol。这说明氨基酸侧链极性的改变增加了侧链与底物分子之间的静电排斥作用,因而静电排斥作用也是导致突变体活性下降的原因之一。同时,突变后系统势能增加了283 kJ/mol,进一步证实了第22位氨基酸侧链极性和带电性质的改变导致酶与底物结合状态的势能增大,从而引起活性大幅下降。因此,将来对酶的结合口袋区域进行理性设计时,除了考虑空间位阻效应外,还需考虑疏水作用和静电作用。Located in the 20s loop of mandelate racemase from Pseudomonas putida,Val22 is one of the amino acids related with substrate binding.In this study,a 75.9% decrease of activity was observed after Val was substituted by Arg.In addition to the reduction of hydrophobic interaction between en-zyme and substrate,electrostatic repulsion could also lead to the decrease of activity.Molecular dy-namic was utilized and showed that the electrostatic potential of substrate and amino acid of No.22 in-creased from 0.036 kJ/mol to 0.124 kJ/mol.This result indicated that the increased activity in the mu-tant was mainly attributed to the shift of side chain polarity and the electrostatic repulsion between side chain and substrate.An increase of 283 kJ/mol in the potential energy of system suggested that the changes of side chain polarity and electric property brought about the increase of potential energy in binding state and the dramatical decline of activity.Therefore hydrophobic interaction and electrostatic interaction should be considered with steric bulk when rationally designing the binding pocket of man-delate racemase.

关 键 词：扁桃酸消旋酶 恶臭假单胞菌 定点突变 分子动力学 

分 类 号：Q936[生物学—微生物学]