The effect of growth hormone and thyroxine on the amount of L-arginine:glycine amidinotransferase in kidneys of hypophysectomized rats. Purification and some properties of rat kidney transamidinase.

The initial step in creatine biosynthesis involves the transfer of an amidine group from arginine to glycine to form guanidinoacetic acid and is catalyzed by the enzyme L-arginine:glycine amidinotransferase (transamidinase). A procedure for the purification of rat kidney transamidinase by chromatography with DEAEcellulose and phenyl-Sepharose is described in this report. Two fractions of enzyme, designated as aand ptransamidinase, were obtained with chromatography on DEAE-cellulose. The two fractions were individually purified to homogeneity as judged by their migration as a single band in native and sodium dodecyl sulfate gel electrophoresis and by sedimentation equilibrium experiments. No significant differences in the properties of the two fractions have been found. The minimum molecular weights, determined by sodium dodecyl sulfate gel electrophoresis, were 42,000 and 44,000. The apparent molecular weights, determined by sedimentation equilibrium, were 82,600 and 83,300. The amino acid compositions and specific activities of the two fractions were similar. The PI values were 6.9 to 7.3. The K,,, values were 2.8 and 2.4 m~ for arginine and 3.0 and 3.1 m~ for glycine. The a and p fractions were identical immunologically. Monospecific antibodies to the purified transamidinase have been prepared. Growth hormone and thyroxine have been reported previously to be necessary for rat kidney transamidinase activities (Van Pilsum, J. F., Carlson, M., Boen, J. R., Taylor, D., and Zakis, B. (1970) Endocrinology 87, 1237-1244). The amounts of transamidinase protein in kidneys from three groups of experimental rats have been determined by immunotitration experiments. Hypophysectomized rats had 30 and 33%. respectively, of the activities and amounts of immunoprecipitable transamidinase protein found in kidneys of sham-hypophysectomized rats. Hypophysectomized rats that had received nine daily injections of growth hormone and thyroxine had similar amounts of transamidinase