The biosynthesis of hyaluronic acid by group A Streptococcus. VI. Biosynthesis from uridine nucleotides in cell-free extracts.

Previous studies have demonstrated that both the glucosamine and glucuronic acid moieties of hyaluronic acid synthesized by Group A streptococci are derived from glucose without previous scission of the glucose molecule (1). Shortly after the discovery by Caputto et al. (2) of the role of uridine diphosphoglucose in the epimerization of glucose, Park (3-5) noted the accumulation of uridine nucleotides in penicillin-inhibited Staphylococcus aureus. Since that time considerable progress has been made in the isolation and characterization of uridine nucleotides containing various carbohydrate moieties. Of particular interest has been the demonstration of uridine diphospho-glucuronate (6)) -N-acetylglucosamine (7-g), and -N-acetylgalactosamine (9). The existence of these compounds, coupled with the demonstration of the role of uridine nucleotides in the biosynthesis of sucrose (lo), sucrose phosphate (1 l), trehalose phosphate (12), and lactose l-phosphate (13), suggested that such compounds are involved in hyaluronate synthesis. Recent demonstrations of the role of uridine nucleotides in chitin (14), cellulose (15), glycogen synthesis (IS), and a p-l ,a-glucan (17) strongly suggest that the donation of glycosyl groups by such nucleotides represents the most general pattern of polysaccharide biosynthesis. Glaser and Brown (18) and Glaser (19) have reported the incorporation of radioactivity from Cl*-labeled uridine diphosphoN-acetylglucosamine into hyaluronate by cell-free extracts of Rous sarcoma. However, the radioactivity was low and could be removed by electrodialysis or reprecipitation of the carrier hyaluronic acid. Previous work has demonstrated that Group A streptococcus Type 18 (strain Alll) contains uridine diphospho-glucose, -glucuronate, and -N-acetylglucosamine (20). The present paper reports the incorporation, by cell-free preparations of streptococci, of radioactivity from tritiated uridine diphospho-l\r-acetylglucosamine and tritiated N-acetylglucosamine l-phosphate into the acetylglucosamine moiety of hyaluronic acid and from tritiated uridine diphosphoglucuronate into the glucuronate moiety of hyaluronic acid. Net synthesis of hyaluronic acid is also demonstrated. A preliminary report of this work has been published (21).