The Structural Bases of Antibiotic Resistance in the Clinically Derived Mutant beta-Lactamases TEM-30, TEM-32, and TEM-34
- NWU
Widespread use of {beta}-lactam antibiotics has promoted the evolution of {beta}-lactamase mutant enzymes that can hydrolyze ever newer classes of these drugs. Among the most pernicious mutants are the inhibitor-resistant TEM {beta}-lactamases (IRTs), which elude mechanism-based inhibitors, such as clavulanate. Despite much research on these IRTs, little is known about the structural bases of their action. This has made it difficult to understand how many of the resistance substitutions act as they often occur far from Ser-130. Here, three IRT structures, TEM-30 (R244S), TEM-32 (M69I/M182T), and TEM-34 (M69V), are determined by x-ray crystallography at 2.00, 1.61, and 1.52 {angstrom}, respectively. In TEM-30, the Arg-244 {yields} Ser substitution (7.8 {angstrom} from Ser-130) displaces a conserved water molecule that usually interacts with the {beta}-lactam C3 carboxylate. In TEM-32, the substitution Met-69 {yields} Ile (10 {angstrom} from Ser-130) appears to distort Ser-70, which in turn causes Ser-130 to adopt a new conformation, moving its O{gamma} further away, 2.3 {angstrom} from where the inhibitor would bind. This substitution also destabilizes the enzyme by 1.3 kcal/mol. The Met-182 {yields} Thr substitution (20 {angstrom} from Ser-130) has no effect on enzyme activity but rather restabilizes the enzyme by 2.9 kcal/mol. In TEM-34, the Met-69 {yields} Val substitution similarly leads to a conformational change in Ser-130, this time causing it to hydrogen bond with Lys-73 and Lys-234. This masks the lone pair electrons of Ser-130 O{gamma}, reducing its nucleophilicity for cross-linking. In these three structures, distant substitutions result in accommodations that converge on the same point of action, the local environment of Ser-130. TEM-1 {beta}-lactamase is the predominant source of resistance to {beta}-lactams, such as the penicillins. TEM-1 and related class A {beta}-lactamases confer resistance by hydrolyzing the {beta}-lactam ring of these antibiotics; bacteria expressing these enzymes have become widespread in hospitals and in the community. Beginning in 1980s, three mechanism-based class A {beta}-lactamase inhibitors, clavulanate, tazobactam, and sulbactam, have been used in combination with conventional penicillins to reverse this resistance (Fig.1, A-C). However, since 1992, more than 26 so-called inhibitor-resistant TEM (IRT)1 mutants have been selected, reversing susceptibility to these three mechanism-based inhibitors in the clinic (www.lahey.org/studies/temtable.stm).
- Research Organization:
- Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Organization:
- USDOE
- OSTI ID:
- 1008803
- Journal Information:
- J. Biol. Chem., Vol. 277, Issue (35) ; 08, 2002; ISSN 0021-9258
- Country of Publication:
- United States
- Language:
- ENGLISH
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