ISOZYMES OF RAT URINARY KALLIKREIN

Two isozymes of rat urinary kallikrein (A and B) have been purified and studied in detail. The enzymes were separated by chromatography on DEAE-cellulose and distinguished by electrophoresis and electrofocusing in polyacrylamide slab gels. The electrophoretic mobilities were not altered by treatment with neuraminidase but were shifted toward the cathode after complexing with aprotinin. Molecular weights of rat urinary kallikreins A and B were estimated to be 36,500 and 35,500, respectively, by sodium dodecyl sulfate polyacrylamide slab gel electrophoresis. The enzymes did not differ in pH optimum, specific activity or substrate specificity, but differed slightly in heat stability. They were not distinguishable biologically, nor immunologically using an antiserum generated in a sheep against kallikrein B. Both kallikreins were activated by deoxycholate and several nonionic detergents. α-N-tosyl-l-arginine methyl ester (Tos-l-Arg OMe) hydrolysis by both forms was inhibited by aprotinin and stimulated by ovomucoid and lima bean trypsin inhibitors, whereas Tos-l-Arg-OMe hydrolysis was stimulated by soybean trypsin inhibitor at low concentrations and inhibited at high concentrations. Kinetic measurements of α-N-benzoyl-l-arginine ethyl ester (Bz-l-Arg-OEt) hydrolysis by the two enzymes showed no significant differences in substrate affinity or in maximal velocity. © 1979.