β-Glucuronidase of Rat Preputial Gland1

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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 crystallization. The purified enzyme appeared homogeneous on electrophoresis in polyacrylamide gel, and on analytical ultracentrifugation and had a molecular weight of approximately 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 0.53 mM. The enzyme had a single pH optimum at 4.5. The enzyme had a very low content of sulphur-containing amino acid and contained 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.

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