Muraki M, Morikawa M, Jigami Y, Tanaka H
National Chemical Laboratory for Industry, Ibaraki, Japan.
Protein Eng. 1988 Apr;2(1):49-54. doi: 10.1093/protein/2.1.49.
The surface positive charges of human lysozyme were either increased or decreased to alter the electrostatic interaction between enzyme and substrate in the lytic action of human lysozyme using site-directed mutagenesis. The amino acid substitutions accompanying either the addition or the removal of two units of positive charge have shifted the optimal ionic strength (NaCl concentration in 10 mM Mes buffer, pH 6.2) for the lysis of Micrococcus lysodeikticus cell from 0.04 M to 0.1 M and from 0.04 M to 0.02 M respectively. In addition to the change in ionic strength-activity profile, the pH-activity profile and the effect of a polycationic electrolyte, poly-L-Lys-HCl, on the lytic activity were significantly changed. Owing to the shifts in both ionic strength profiles and pH profiles the Arg74/Arg126 mutant has become a better catalyst than wild-type enzyme under the conditions of high ionic strength and high pH, and the Gln41/Ser101 mutant has become a better catalyst under the conditions of low ionic strength and low pH.
利用定点诱变技术改变人溶菌酶的表面正电荷，以改变人溶菌酶在裂解作用中酶与底物之间的静电相互作用。伴随正电荷增加或减少两个单位的氨基酸替换，分别使溶壁微球菌细胞裂解的最佳离子强度（10 mM Mes缓冲液，pH 6.2中的NaCl浓度）从0.04 M变为0.1 M以及从0.04 M变为0.02 M。除了离子强度-活性曲线的变化外，pH-活性曲线以及聚阳离子电解质聚-L-赖氨酸盐酸盐对裂解活性的影响也发生了显著变化。由于离子强度曲线和pH曲线的变化，在高离子强度和高pH条件下，Arg74/Arg126突变体已成为比野生型酶更好的催化剂，而在低离子强度和低pH条件下，Gln41/Ser101突变体已成为更好的催化剂。
