Kinetics and stereochemistry of hydrolysis of an N-(phenylacetyl)--hydroxyglycine ester catalyzed by serine -lactamases and dd-peptidasesElectronic supplementary information (ESI) available: NMR spectra of 5 and its reaction products, and HPLC experiments. See DOI: 10.1039/c2ob25585e

• Main Authors: Pelto, Ryan B, Pratt, R. F
• Format: Article
• Language: English
• ISSN: 1477-0520; 1477-0539
• DOI: 10.1039/c2ob25585e

The -hydroxydepsipeptide 3-carboxyphenyl N -(phenylacetyl)--hydroxyglycinate ( 5 ) is a quite effective substrate of serine -lactamases and low molecular mass dd -peptidases. The class C P99 and ampC -lactamases catalyze the hydrolysis of both enantiomers of 5 , although they show a strong preference for one of them. The class A TEM-2 and class D OXA-1 -lactamases and the Streptomyces R61 and Actinomadura R39 dd -peptidases catalyze hydrolysis of only one enantiomer of 5 at any significant rate. Experiments show that all of the above enzymes strongly prefer the same enantiomer, a surprising result since -lactamases usually prefer l ( S ) enantiomers and dd -peptidases d ( R ). Product analysis, employing peptidylglycine -amidating lyase, showed that the preferred enantiomer is d ( R ). Thus, it is the -lactamases that have switched preference rather than the dd -peptidases. Molecular modeling of the P99 -lactamase active site suggests that the -hydroxyl of 5 may interact with conserved Asn and Lys residues. Both -hydroxy and -amido substituents on a glycine ester substrate can therefore enhance its productive interaction with the -lactamase active site, although their effects are not additive; this may also be true for inhibitors. -Lactam-recognizing enzymes make a choice between an R and an S -hydroxyl group.