Fish muscle riboside hydrolases

Abstract

A method of preparation of crude riboside hydrolase enzymes from muscles of certain marine fishes is described. The muscle is extracted with cold 0.5 - 0.6 [image] NaCl or KC1 at pH 4.6 and the extract fractionated with (NH4)2SO4 at pH 7, the fraction obtained between 0.4 and 0.6 saturation possessing most of the activity. Crude preparations were purified by stirring with A000mberlite cation-exchange resin XE 64 at pH values slightly above 7, eluting with NaCl and di-alyzing. Crude preparations contained 2 (and occasionally 3) proteins as judged by zone electrophoresis while purified preparations showed a single protein zone. The purest preparation obtained hydrolyzed guanosine over 1500 times as actively as did whole muscle on a protein N basis. Purified preparations contained 2 distinct enzymes, a non-specific riboside hydrolase with a pH optimum of about 5.5 which hydrolyzed purine ribosides and cytidine, and a specific inosine hydrolase with a pH optimum of about 8.0. These enzymes were not separated. Neither ribose 1-phosphate nor ribose 5-phosphate was hydrolyzed, and nuceloside hydrolysis proceeded in the absence of added inorganic phosphate. D-Ribose diphenylhydrazone and xanthine were isolated from a reaction mixture of guanosine and crude riboside hydrolase. The enzyme preparations withstood freeze-drying and were active even at 55[degree].