Deacylation Kinetics Analysis of Streptococcus pneumoniae Penicillin-Binding Protein 2x Mutants Resistant to β-Lactam Antibiotics Using Electrospray Ionization– Mass Spectrometry
✍ Scribed by Anne Marie Di Guilmi; Nicolas Mouz; Yves Pétillot; Eric Forest; Otto Dideberg; Thierry Vernet
- Publisher
- Elsevier Science
- Year
- 2000
- Tongue
- English
- Weight
- 115 KB
- Volume
- 284
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
Penicillin-binding proteins (PBPs) catalyze the transpeptidase reaction involved in peptidoglycan synthesis and are covalently inhibited by the -lactam antibiotics. In a previous work we have focused on acylation efficiency measurements of various Streptococcus pneumoniae PBP2x* mutants to study the molecular determinants of resistance to -lactams. In the present paper we have developed a method to improve an accurate determination of the deacylation rate constant using electrospray ionization-mass spectrometry. This method is adaptable to the analysis of deacylation of any -lactam. Compared to the fluorographic technique, the ESI-MS method is insensitive to variations in the concentration of functional proteins and is therefore more reliable. We have established that the resistance of PBPs to -lactams is mostly due to a decrease of the acylation efficiency with only marginal effects on the deacylation rates.