Determination of murein precursors during the cell cycle of Escherichia coli

Abstract

A convenient and reliable method has been established that allows a quantitative detemination of m-diamino[3H]pimelic acid-labelled murein precursors in 1 ml culture samples of Escherichia coli. Prior to separation by reversed-phase high-pressure liquid chromatography the lipid-linked intermediates were hydrolysed to release the muropeptides. The accuracy for the measurement of UDP-N-acetylmuramylpentapeptide (UDP-MurNAc-pentapeptide) was ± 1.9% (SD), for undecaprenyl-P-P-MurNAc--pentapeptide (lipid I) ± 10% (SD) and for undecaprenyl-P-P-(GlcNAc-β1→4)MurNAc-pentapeptide (lipid II) ± 5% (SD). The ratio of UDP-MurNAc-pentapeptide: lipid I: lipid II was aobut 300:1:3 for E. coli MC4100. The relative cellular concentrations of all three precursor molecules were found not to vary throughout the cell cycle. It is concluded that elongation and division of the murein saculus is not controlled by oscillations in the concentrations of these late murein precursors.