Abstract
Bacterial cell wall peptidoglycan is synthesized from lipid II precursor by two reactions. Glycosyltransferases polymerize the glycan chains and transpeptidases form the peptide cross-links. The bifunctional class A penicillin-binding proteins catalyze both of these reactions. Here, we describe an in vitro peptidoglycan synthesis assay utilizing radiolabeled lipid II substrate to monitor simultaneously peptidoglycan glycosyltransferase and transpeptidase activities. The products of the reaction are separated by high-pressure liquid chromatography and quantified by flow-through scintillation counting.
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Acknowledgements
We thank Carolin Ewers for the SDS-PAGE analysis shown in Fig. 1b. This work was supported by the Biotechnology and Biological Sciences Research Council (BBSRC) and the EU Commission within the DIVINOCELL and EUR-INTAFAR projects.
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Biboy, J., Bui, N.K., Vollmer, W. (2013). In Vitro Peptidoglycan Synthesis Assay with Lipid II Substrate. In: Delcour, A. (eds) Bacterial Cell Surfaces. Methods in Molecular Biology, vol 966. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-245-2_17
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DOI: https://doi.org/10.1007/978-1-62703-245-2_17
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