Human Placental N-Acetyl-/?-o-Hexosaminidase lsozymes Activity toward Native Hyaluronic Acid

The activity of purified human hexosaminidases A and B toward hyaluronic acid (HA) isolated from cultured hum& skin fibroblasts was investigated. The cleavage of N-acetylglucosaminyl residues to monosaccharide N-acetylglucosamines by hexosaminidase isozymes was determined in the presence and absence of purified human fi-glucuronidase. The pH optima of this reaction, with and without P-glucuronidase, were 4.5 for hexosaminidase A and 4.0 for hexosaminidase B. The hydrolysis of HA by both hexosaminidase isozymes proceeds linearily for at least 18 h in the presence of /3-glucuronidase. Concentrations of 0.5-5 units of either isozyme showed a linear relationship with rate of hydrolysis. Without /3-glucuronidase, hexosaminidase only cleaved the terminal N-acetylglucosamine residue. However, under optimal conditions, with j?-glucuronidase, the hydrolytic activity of hexosaminidase B was about 30% as efficient as that of hexosaminidase A. Approximately 70% of the HA could be degraded by 5 units of hexosaminidase A in the presence of 0.5 unit of P-glucuronidase, as opposed to 25% degraded by hexosaminidase B. These results probably reflect intrinsic differences in the activities of the two isozymes. Since the substrate (HA) did not inhibit the hydrolysis of a synthetic substrate (4-methylumbelliferyl-/?-glucosaminide) by hexosaminidase B, the linear kinetics of HA hydrolysis implies no product inhibition. These data indicate that native HA can be hydrolyzed by the combined activities of /3glucuronidase with hexosaminidase A or hexoaminidase B.