Peptidoglycan synthesis by partly autolyzed cells of Bacillus subtilis W23.
Partly autolyzed, osmotically stabilized cells of Bacillus subtilis W23 synthesized peptidoglycan from the exogenously supplied nucleotide precursors UDP-N-acetylglucosamine and UDP-N-acetylmuramyl pentapeptide. Freshly harvested cells did not synthesize peptidoglycan. The peptidoglycan formed was entirely hydrolyzed by N-acetylmuramoylhydrolase, and its synthesis was inhibited by the antibiotics bacitracin, vancomycin, and tunicamycin. Peptidoglycan formation was optimal at 37 degrees C and pH 8.5, and the specific activity of 7.0 nmol of N-acetylglucosamine incorporated per mg of membrane protein per h at pH 7.5 was probably decreased by the action of endogenous wall autolysins. No cross-linked peptidoglycan was formed. In addition, a lysozyme-resistant polymer was also formed from UDP-N-acetylglucosamine alone. Peptidoglycan synthesis was inhibited by trypsin and p-chloromercuribenzenesulfonic acid, and we conclude that it occurred at the outer surface of the membrane. Although phospho-N-acetylmuramyl pentapeptide translocase activity was detected on the outside surface of the membrane, no transphosphorylation mechanism was observed for the translocation of UDP-N-acetylglucosamine. Peptidoglycan was similarly formed with partly autolyzed preparations of B. subtilis NCIB 3610, B. subtilis 168, B. megaterium KM, and B. licheniformis ATCC 9945. Intact protoplasts of B. subtilis W23 did not synthesize peptidoglycan from externally supplied nucleotides although the lipid intermediate was formed which was inhibited by tunicamycin and bacitracin. It was therefore considered that the lipid cycle had been completed, and the absence of peptidoglycan synthesis was believed to be due to the presence of lysozyme adhering to the protoplast membrane. The significance of these results and similar observations for teichoic acid synthesis (Bertram et al., J. Bacteriol. 148:406-412, 1981) is discussed in relation to the translocation of bacterial cell wall polymers.
D001412:Bacillus subtilis / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects; D050356:Lipid Metabolism; D009713:Nucleotidyltransferases / Q000378:metabolism; D010457:Peptidoglycan / Q000096:biosynthesis*; D011523:Protoplasts; D014050:Toluene / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology
D010457:Peptidoglycan; D014050:Toluene; D009713:Nucleotidyltransferases; C023226:UDPacetylglucosamine pyrophosphorylase; D014357:Trypsin