Biosynthesis of the Polysaccharide of Micrococcus lysodeikticus Cell Walls

The particulate enzyme fraction obtained from Micrococcus lysodeikticus catalyzed the incorporation of d-[14C]glucose from uridine diphospho-d-[14C]glucose into a polymer which resembled the d-glucose- and N-acetyl-d-mannosaminuronic acid-containing polysaccharide of the cell walls of this organism. In vitro biosynthesis of the polysaccharide depended upon the presence of the uridine diphospho-N-acetylhexosaminuronic acid fraction isolated from M. lysodeikticus and uridine diphospho-N-acetyl-d-glucosamine and was stimulated by a heat-stable factor obtained from the soluble fraction of cell extracts. The reaction proceeded optimally at pH 8.2 and at 20 mm magnesium ion concentration. The incorporation of d-[14C]glucose proceeded after a lag period of 15 to 20 min. Preincubation of the reaction components with the particulate enzyme fraction before the addition of uridine diphospho-d-glucose eliminated the lag period. [14C]N-Acetylhexosaminuronic acid residues were incorporated into polysaccharide from [14C]uridine diphospho-N-acetylhexosaminuronic acid (labeled throughout the molecule) in an amount approximately equimolar to d-glucose in a reaction dependent upon uridine diphospho-d-glucose. However, when uridine diphospho-N-[14C]acetyl-d-glucosamine was the labeled substrate, N-[14C]acetyl-d-glucosamine was incorporated only to the extent of 1 residue or less for each 15 residues of d-glucose.