Insulin-like activity of proteases. V. Stimulation of cyclic adenosine 3',5'-monophosphate phosphodiesterase by an N-succinyl-L-alanyl-L-alanyl-L-alanine p-nitroanilide-hydrolyzing protease.

Abstract

The stimulatory effect of an N-succinyl-L-alanyl-L-alanyl-L-alanine p-nitroanilidehydrolyzing protease (STA-protease), which was partially purified from Pronase, on cyclic adenosine 3', 5'-monophosphate (cyclic AMP) phosphodiesterase of bovine heart was investigated by measuring the changes in kinetic constants. The phosphodiesterase used showed hydrolytic activity towards cyclic AMP at a concentration of 1 or 100 μM. STA-protease stimulated low Km phosphodiesterase but not high Km phosphodiesterase activity, in the presence of ethylene glycol bis (β-aminoethylether)-N, N'-tetraacetic acid (EGTA). No stimulation of the low Km enzyme was observed when theophylline was used instead of EGTA. Proteases such as Pronase, trypsin, chymotrypsin, elastase, and substilisin BPN' stimulated the low Km enzyme to various extents. Although STA-protease showed the lowest caseinolytic activity among proteases tested, its stimulatory activity towards the low Km enzyme was rather high. STA-protease increased the Km and Vmax values of the low Km enzyme, whereas trypsin decreased the Km value without causing significant change in Vmax. The stimulatory mechanism of STA-protease probably differs from that of trypsin.