Structural milestones in the reaction pathway of an amide hydrolase: Substrate, acyl, and product complexes of cephalothin with AmpC β-lactamase

Beth M. Beadle, Indi Trehan, Pamela J. Focia, Brian K. Shoichet*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

100 Scopus citations

Abstract

β-lactamases hydrolyze β-lactam antibiotics and are the leading cause of bacterial resistance to these drugs. Although β-lactamases have been extensively studied, structures of the substrate-enzyme and product-enzyme complexes have proven elusive. Here, the structure of a mutant AmpC in complex with the β-lactam cephalothin in its substrate and product forms was determined by X-ray crystallography to 1.53 Å resolution. The acyl-enzyme intermediate between AmpC and cephalothin was determined to 2.06 Å resolution. The ligand undergoes a dramatic conformational change as the reaction progresses, with the characteristic six-membered dihydrothiazine ring of cephalothin rotating by 109°. These structures correspond to all three intermediates along the reaction path and provide insight into substrate recognition, catalysis, and product expulsion.

Original languageEnglish (US)
Pages (from-to)413-424
Number of pages12
JournalStructure
Volume10
Issue number3
DOIs
StatePublished - 2002

Keywords

  • AmpC
  • Cephalothin
  • Product-enzyme complex
  • Substrate-enzyme complex
  • X-ray crystallography
  • β-lactamase

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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