β-Aspartylglucosylamine Amido Hydrolase of Rat Liver and Kidney

Abstract

A variety of rat tissues have the ability to hydrolyze 1-l-β-aspartamido[-2-acetamido]-1,2-dideoxy-β-d-glucose. In rat liver and kidney the enzyme is present in lysosomes. The lysosomal nature of the enzyme was established by the amount of enzyme sedimentable with mitochondrial and light mitochondrial particles, by the purification of lysosomes, and by the isolation of Triton WR 1339-filled liver lysosomes. Freezing and thawing, osmotic shock, and treatment with Triton X-100 resulted in the release of the enzyme from lysosomes. The kidney lysosomal enzyme was purified 430-fold by Sephadex gel filtration and the liver enzyme 650-fold by chromatography on carboxymethyl cellulose. In both instances the enzyme was well separated from β-N-acetylglucosaminidase. The pH optimum of the enzyme was 7.6. The K_m was 5.9 x 10^-4 m. The enzyme was inhibited by p-chloromercuribenzoate, and by copper and nickel ions. Other divalent metal ions did not have any effect. The enzyme was quite stable in dilute solutions for prolonged periods of time. The purified enzyme had no asparaginase activity, but liberated aspartic acid from the substrate. It is concluded that the enzyme is an amidase. It is suggested that the amidase is involved in the intracellular digestion of glycoproteins.