Myocardial guanylate cyclase: Properties of the enzyme and effects of cholinergic agonists in vitro

Abstract

The characteristics of myocardial guanylate cyclase (GTP pyrophosphatelyase, EC 4.6.1.2) were studied. Specific activity of the myocardial enzyme in five vertebrate species was guinea pig > man ⪢ cat > dog > rat. In the guinea pig, guanylate cyclase activity was uniformly distributed throughout the anatomical regions of the heart. The major portion of the enzyme activity was retrieved in the supernatant fraction after centrifugation at 12 000 × g. The K m for GTP was similar in supernatant (0.12 mM) and particulate (0.21 mM) preparations, although the K a for Mn 2+ in particulate preparations (0.3–0.6 mM) was less than that observed for guanylate cyclase in the supernatant fraction (0.8–2.0 mM). ATP competitively inhibited supernatant and particulate activity. Addition of 0.005–10.0 mM Ca 2+ to assay incubations did not enhance guanylate cyclase activity. Suspension of 105 000 × g supernatant guanylate cyclase preparations with membrane lipids or phosphatidylserine stimulated activity 1.4–4.3 fold, whereas similar treatment of particulate preparations caused little alteration of enzyme activity. Addition of the cholinergic agonists acetylcholine, carbachol or methacholine (10 −4–10 −8 M) to homogenate, supernatant, particulate and disrupted tissue slice preparations in the presence of 0.0012–1.2 mM GTP, 0.3–10.0 mM Mn 2+ and 0.005–10.0 mM Ca 2+ or 0.0012–1.2 mM ATP did not stimulate guanylate cyclase activity. Similarly, further stimulation of guanylate cyclase activity was not elicited when enzym-lipid suspensions were assayed in the presence of cholinergic agents.