Molecular Forms of Rat-Liver Arginase. Isolation and Characterization.

Debate continues over the physical characteristics and even the existence of arginase isoenzymes. This paper gives additional support for such multiplicity and reports differences in physical characteristics among the various forms. After 2500-5000-fold purification of rat liver arginase, three molecular forms were separated on carboxymethyl-cellulose columns and were purified 2500-5000-fold, 800-1000-fold and 600-1000-fold, respectively. The molecular forms have also been identified by chromatography in the supernatant of tissue extracts. The isolation of these molecular forms by affinity chromatography, using Sepharose-lysine as a competitive inhibitor of arginase, shows only one main form, however. Kinetic studies were done for two of the molecular forms isolated, specifically the activation energy (Ea, the energy of denaturatization (Ed), Km, pH and the effect of divalent cations were determined. Significant differences were found for the Ea between the two molecular forms. The isolated isoenzymes are cationic at pH 5.5 and pH 8.8. However, they show different mobilities in electrophoresis. The molecular weight determination by gel filtration yields a value of 110000 to 115000 for both forms. The use of thin-layer immunochromatography plates, a combination of molecular weight and immunodiffusion technique, gave only one peak with the same molecular weight as that determined by gel filtration. The immunological studies showed that the isoenzymes have similar antigenic determinants. [ABSTRACT FROM AUTHOR]