Structural analysis of Bacillus megaterium KM spore peptidoglycan and its dynamics during germination

The composition and structure of peptidoglycan from dormant spores of Bacil lus megaterium MM and its dynamics during germination were investigated. Am ino acid analysis and mass spectrometry identified 21 muropeptides resolved by reverse phase HPLC following digestion of peptidoglycan with Cellosyl. The basic structure of peptidoglycan in B. megaterium spores is similar to that of Bacillus subtilis: 44.2 % of muramic acid residues are substituted with delta-lactam, 28.8% with single L-alanine, 25.1 % with tetrapeptide an d only 1.8% with tripeptide. The cross-linking index of the spore peptidogl ycan, determined from muropeptides resolved by reverse phase HPLC, was 2.2 % per muramic acid. Spore peptidoglycan contains 2.9 % of muropeptides with unsubstituted N-acetylmuramic acid. These muropeptides are likely to be in termediate products of delta-lactam formation. Analysis of muropeptide dyna mics during germination revealed the activity of at least two hydrolytic en zymes, an N-acetylglucosaminidase and a lytic transglycosylase. A 4 M LiCl extract from 30 min germinated spores of B. megaterium KM caused 'germinati on-like' changes to permeabilized spores of B. megaterium and B. subtilis b ut not those of a B. subtilis cwlD mutant. Muropeptide analysis of the trea ted spores revealed the presence of products generated by the activity of a glucosaminidase.