Inhibition of peptidoglycan biosynthesis at a postcytoplasmic reaction in a stable L-phase variant of Streptococcus faecium.

Cultures of a stable L-phase variant of Streptococcus faecium F24 produced and retained peptidoglycan precursors intracellularly over the entire growth cycle in a chemically defined medium. The identity of the most abundant precursor, UDP N-acetylmuramyl-L-alanyl-D-glutamyl-L-lysyl-D-alanyl-D-alanine (UDP-MurNAc-pentapeptide), was confirmed by demonstrating in vitro the presence of enzymes required for the cytoplasmic stage of peptidoglycan biosynthesis. The initial membrane-bound reaction in peptidoglycan biosynthesis involving phospho-MurNAc-pentapeptide translocase and undecaprenyl-phosphate membrane carrier was catalyzed by protoplast membrane preparations but not by L-phase membrane preparations. However, both protoplast and L-phase membranes incorporated radioactivity from dTDP-L-[14C]rhamnose, the presumed precursor to a non-peptidoglycan cell surface component, into high-molecular-weight material. dTDP-L-rhamnose did not accumulate in growing cultures but was synthesized from D-glucose-1-phosphate and dTTP by cell-free extracts of the streptococcus and L-phase variant. Neither rhamnose- nor muramic acid-containing compounds were detected in culture fluids. It is suggested that continued inhibition of cell wall biosynthesis in this stable L-phase variant is the result of a defect expressed at the membrane stage of peptidoglycan biosynthesis specifically involving the translocation step.