Abstract
Rat preputial gland β-glucuronidase [EC 3.2.1. 31] was purified by ammonium sulfate precipitation, ethanol fractionation, gel filtration on Sephadex G-200 and crystalliza-tion.
The purified enzyme appeared homogeneous on electrophoresis in polyacrylamide gel, and on analytical ultracentrifugation and had a molecular weight of approxi-mately 320, 000, and a sedimentation coefficient of 12S. SDS polyacrylamide gel electrophoresis indicated that the enzyme consisted of subunits with molecular weights of 79, 000, so the native enzyme appeared to be a tetramer. The Km with p-nitrophenyl β-D-glucosiduronic acid as substrate was about 0.53mM. The enzyme had a single pH optimum at 4.5.
The enzyme had a very low content of sulphur-containing amino acid and con-tained 5.7% carbohydrate, consisting of mannose, glucose, fucose, galactose, and glucosamine in a ratio of 44; 9; 6; 2; 41. Sialic acid was not detected in the crystallized enzyme.
