Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0276640 (TEM)
 20,729 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Beta-lactamase acquisition is the most prevalent basis for Gram-negative bacteria resistance to the beta-lactam antibiotics. The mechanism used by the most common class A Gram-negative beta-lactamases is serine acylation followed by hydrolytic deacylation, destroying the beta-lactam. The ab initio quantum mechanical/molecular mechanical (QM/MM) calculations, augmented by extensive molecular dynamics simulations reported herein, describe the serine acylation mechanism for the class A TEM-1 beta-lactamase with penicillanic acid as substrate. Potential energy surfaces (based on approximately 350 MP2/6-31+G calculations) reveal the proton movements that govern Ser70 tetrahedral formation and then collapse to the acyl-enzyme. A remarkable duality of mechanism for tetrahedral formation is implicated. Following substrate binding, the pathway initiates by a low energy barrier (5 kcal mol(-1)) and an energetically favorable transfer of a proton from Lys73 to Glu166, through the catalytic water molecule and Ser70. This gives unprotonated Lys73 and protonated Glu166. Tetrahedral formation ensues in a concerted general base process, with Lys73 promoting Ser70 addition to the beta-lactam carbonyl. Moreover, the three-dimensional potential energy surface also shows that the previously proposed pathway, involving Glu166 as the general base promoting Ser70 through a conserved water molecule, exists in competition with the Lys73 process. The existence of two routes to the tetrahedral species is fully consistent with experimental data for mutant variants of the TEM beta-lactamase.
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PMID:Ab initio QM/MM study of class A beta-lactamase acylation: dual participation of Glu166 and Lys73 in a concerted base promotion of Ser70. 1626 3

The breakdown of beta-lactam antibiotics by beta-lactamases is the most important resistance mechanism of gram negative bacteria against these drugs. The reaction mechanism of class A beta-lactamases, the most widespread family of these enzymes, consists of two main steps: acylation of an active site serine by the antibiotic, followed by deacylation and release of the cleaved compound. We have investigated the first step in acylation (the formation of the tetrahedral intermediate) for the reaction of benzylpenicillin in the TEM-1 enzyme using high level combined quantum mechanics/molecular mechanics (QM/MM) methods. Structures were optimized at the B3LYP/6-31+G(d)/CHARMM27 level, with energies for key points calculated up to the ab initio SCS-MP2/aug-cc-pVTZ/CHARMM27 level. The results support a mechanism in which Glu166 removes a proton (via an intervening water molecule) from Ser70, which in turn attacks the beta-lactam of the antibiotic. Depending on the method used, the calculated barriers range from 3 to 12 kcal mol(-1) for this step, consistent with experimental data. We have also modeled this reaction step in a model of the K73A mutant enzyme. The barrier to reaction in this mutant model is found to be slightly higher: the results indicate that Lys73 stabilizes the transition state, in particular deprotonated Ser70, lowering the barrier by about 1.7 kcal mol(-1). This finding may help to explain the conservation of Lys73, in addition to the role we have previously found for it in the later stages of the reaction (Hermann et al. Org. Biomol. Chem. 2006, 4, 206-210).
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PMID:High level QM/MM modeling of the formation of the tetrahedral intermediate in the acylation of wild type and K73A mutant TEM-1 class A beta-lactamase. 1979 86