Isolation and Characterization of Structural Components of Bacillus cereus AHU 1356 Cell Walls

From lysozyme digests of the N-acetylated cell walls of Bacillus cereus AHU 1356, a polysaccharide fraction and a teichoic acid fraction were isolated by ion-exchange chromatography and gel chromatography. The former fraction, accounting for 50% of the walls, contained N-acetylglucosamine, N-acetylmannosamine, N-acetylgalactosamine and glucose in a molar ratio of 4:1:1:1 together with a small amount of the peptidoglycan constituents. The latter fraction accounted for 5% of the walls and was composed of N-acetylglucosamine, galactose, glycerol and phosphorus in a molar ratio of 1:1.4:1:1 and a small amount of the peptidoglycan constituents. The molecular weight of the polysaccharide fraction was about 33000 as estimated by gel chromatography. After acid hydrolysis of the polysaccharide fraction, a stoichiometric amount of muramic acid 6-phosphate was detected which was probably involved in the linkage between the polysaccharide and peptidoglycan moieties. The polysaccharide moiety detached from the peptidoglycan components by mild acid hydrolysis was isolated as material with an approximate molecular weight of 28000. Structural studies on the polysaccharide, involving Smith degradation, CrO3 oxidation and proton magnetic resonance spectrometry of the polysaccharide fraction together with the analysis of oligosaccharides obtained from partial acid hydrolysis of the polysaccharide fraction, led to the tentative formulation of the most likely structure of the repeating unit as follows: