Continuous Fluorescence Assay for Peptidoglycan Glycosyltransferases.
Bacterial Cell Wall Homeostasis
Methods in molecular biology, January 2016
Alexander J. F. Egan, Waldemar Vollmer
Bacterial cell wall peptidoglycan is synthesized from its precursor lipid II by two enzymatic reactions. First, glycosyltransferases polymerize the glycan strands and second, DD-transpeptidases form cross-links between peptides of neighboring strands. Most bacteria possess bifunctional peptidoglycan synthesis enzymes capable of catalyzing both reactions. Here, we describe a continuous fluorescence glycosyltransferase assay using Dansyl-labeled lipid II as substrate. Progression of the reaction is monitored by the reduction in fluorescence over time. The assay is suitable to investigate the effect of protein interaction partners on the glycan strand synthesis activity of peptidoglycan polymerases.
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