pK(a) calculations for class A beta-lactamases: Methodological and mechanistic implications

被引:57
作者
Raquet, X
Lounnas, V
LamotteBrasseur, J
Frere, JM
Wade, RC
机构
[1] EUROPEAN MOL BIOL LAB, D-69117 HEIDELBERG, GERMANY
[2] UNIV LIEGE, CTR INGN PROT, B-4000 LIEGE, BELGIUM
关键词
D O I
10.1016/S0006-3495(97)78270-5
中图分类号
Q6 [生物物理学];
学科分类号
071011 ;
摘要
beta-lactamases are responsible for resistance to penicillins and related beta-lactam compounds. Despite numerous studies, the identity of the general base involved in the acylation step is still unclear. It has been proposed, on the basis of a previous pK(a) calculation and analysis of structural data, that the unprotonated Lys(73) in the active site could act as the general base. Using a continuum electrostatic model with an improved treatment of the multiple titration site problem, we calculated the pK(a) values of all titratable residues in the substrate-free TEM-1 and Bacillus licheniformis class A beta-lactamases. The pK(a) of Lys(73) in both enzymes was computed to be above 10, in good agreement with recent experimental data on the TEM-1 beta-lactamase, but inconsistent with the proposal that Lys(73) acts as the general base. Even when the closest titratable residue, Glu(166), is mutated to a neutral residue, the predicted downward shift of the pK(a) of Lys(73) shows that it is unlikely to act as a proton abstractor in either enzyme. These results support a mechanism in which the proton of the active Ser(70) is transferred to the carboxylate group of Glu(166).
引用
收藏
页码:2416 / 2426
页数:11
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