Tsukamoto K, Nishida N, Tsuruoka M, Sawai T
Division of Microbial Chemistry, Faculty of Pharmaceutical Sciences, Chiba University, Japan.
FEBS Lett. 1990 Oct 1;271(1-2):243-6. doi: 10.1016/0014-5793(90)80416-g.
The conserved KTG triad in the class C beta-lactamase from Citrobacter freundii GN346 was examined as to its function by means of site-directed mutagenesis. The following conversions were performed; Lys-315 to arginine, alanine or glutamic acid, Thr-316 to valine, and Gly-317 to alanine, proline or isoleucine. The resultant mutant enzymes revealed that a basic amino acid at position 315 and a small uncharged residue at position 317 are essential for the enzyme activity, but a hydroxyl group at residue 316 is not required for the enzymatic catalysis. The kinetic properties of the purified Arg-315 and Val-316 enzymes provided information on the function of these residues.
通过定点诱变研究了弗氏柠檬酸杆菌GN346的C类β-内酰胺酶中保守的KTG三联体的功能。进行了以下替换：将赖氨酸-315替换为精氨酸、丙氨酸或谷氨酸，苏氨酸-316替换为缬氨酸，甘氨酸-317替换为丙氨酸、脯氨酸或异亮氨酸。所得突变酶表明，315位的碱性氨基酸和317位的小的不带电荷的残基对酶活性至关重要，但316位残基上的羟基对于酶催化不是必需的。纯化的精氨酸-315和缬氨酸-316酶的动力学性质提供了关于这些残基功能的信息。
