Gly-238-Ser substitution changes the substrate specificity of the SHV class A β-lactamases

K. Y. Lee, J. D. Hopkins, T. F. O'Brien, M. Syvanen

Research output: Contribution to journalArticle

26 Scopus citations

Abstract

The SHV-type β-lactamase SHV-2A is related to SHV-1 by a Gly-238-Ser replacement. Strains carrying SHV-2A are resistant to the third generation cephems cefotaxime and ceftizoxime, whereas those that carry SHV-1 are sensitive to these drugs. We present a kinetic analysis of a SHV-1 and SHV-2A enzymes, with the goal of gaining insight into the role of residue 238 in hydrolyzing cefotaxime and ceftizoxime. SHV-2A shows altered kinetic properties for a number of other cephems that also have heterocyclic side chains at the amino position of the 7-aminocephalosporanic acid nucleus (R1 side chain), including a significantly higher k(cat)/K(m) than does SHV-1 for cephaloridine, cephalothin, and cefotiam. Two cephems with straight chain R1 substitutions, cephalosporin C and cephacetrile, are not hydrolyzed more efficiently by SHV-2A. These results indicate that the Ser-238-Gly substitution increases the affinity toward cephems with a heterocyclic ring in the R1 side chain. In addition, the data for ampicillin and benzylpenicillin show that addition of a nitrogen to the second carbon of the R1 side chain of a penem results in a lower k(cat)/K(m) for SHV-2A relative to SHV-1. These data strongly suggest that the previously proposed hydrogen bond formation between Ser-238 and the second carbon nitrogen of cefotaxime is not an important factor in hydrolysis by SHV-2A. We propose that the Gly-238 to Ser-238 replacement in SHV-2A has altered the hydrophobic pocket so that it can better accommodate cephems with bulky R1 side chains.

Original languageEnglish (US)
Pages (from-to)45-51
Number of pages7
JournalProteins: Structure, Function and Genetics
Volume11
Issue number1
StatePublished - 1991

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Keywords

  • cefotaxime resistance
  • enzyme structure-function
  • third generation cephalosporins

ASJC Scopus subject areas

  • Genetics
  • Structural Biology
  • Biochemistry

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