Separation and properties of two arylamidases from rat cardiac-muscle extracts

Two main arylamidase activities were separated from a particle-free supernatant of rat heart by chromatography on DEAE-Sephadex. Although both Enzymes hydrolysed L-leucine 4-nitroanilide, only peak-II Enzyme hydrolysed L-lysine 4-nitroanilide. A third minor peak (Ia) contained an Enzyme that was active mainly on the L-lysine 4-nitroanilide. The mol.wts. of the Enzymes in peaks I and II were approx. 257000 and 105000 respectively. The pH optimum was approx. pH7.0 for peak-I Enzyme and 7.0-8.0 for peak-II Enzyme. Both Enzymes were inhibited by addition of puromycin, p-hydroxymercuribenzoate, o-phenanthroline and bivalent metal ions. Addition of dithiothreitol resulted in stimulation of both activities. Dialysis against o-phenanthroline resulted in inhibition of peak-I and -II Enzymes, but after dialysis against EDTA only peak-II Enzyme was inhibited.