The structural bases of antibiotic resistance in the clinically derived mutant β-lactamases TEM-30, TEM-32, and TEM-34

Xiaojun Wang, George Minasov, Brian K. Shoichet*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

61 Scopus citations

Abstract

Widespread use of β-lactam antibiotics has promoted the evolution of β-lactamase mutant enzymes that can hydrolyze ever newer classes of these drugs. Among the most pernicious mutants are the inhibitor-resistant TEM β-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 (M691/MI82T), and TEM-34 (M69V), are determined by x-ray crystallography at 2.00, 1.61, and 1.52 A,respectively. In TEM-30, the Arg-244 ←Ser substitution (7.8 A from Ser-130) displaces a conserved water molecule that usually interacts with the β-lactam C3 carboxylate. In TEM-32, the substitution Met-69 - ← Ile (10 A from Ser-130) appears to distort Ser-70, which in turn causes Ser-130 to adopt a new conformation, moving its Oy further away, 2.3 A from where the inhibitor would bind. This substitution also destabilizes the enzyme by 1.3 kcal/mol. The Met-182 ← Thr substitution (20 A 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 ← 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 Oy, 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.

Original languageEnglish (US)
Pages (from-to)32149-32156
Number of pages8
JournalJournal of Biological Chemistry
Volume277
Issue number35
DOIs
StatePublished - Aug 30 2002

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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