title: Calcium ion downregulates soluble guanylyl cyclase activity: evidence for a two-metal ion catalytic mechanism.
abstract: The inhibition of soluble guanylyl cyclase by Ca2+ has been kinetically characterized and the results support a two-metal-ion catalytic mechanism for formation of cGMP. Ca2+ reversibly inhibits both the basal and NO-stimulated forms of bovine lung soluble guanylyl cyclase. Inhibition is independent of the activator identity and concentration, revealing that Ca2+ interacts with a site independent of the heme regulatory site. Inhibition by Ca2+ is competitive with respect to Mg2+ in excess of substrate, with Kis values of 29 +/- 4 and 6.6 +/- 0.6 microM for the basal and activated states, respectively. Ca2+ inhibits noncompetitively with respect to the substrate MgGTP in both activity states. The qualitatively similar inhibition pattern and quantitatively different Ki values between the basal and NO-stimulated states suggest that the Ca2+ binding site undergoes some structural modification upon activation of the enzyme. The competitive nature of Ca2+ inhibition with respect to excess Mg2+ is consistent with a two-metal-ion mechanism for cyclization.
authors.affiliation: Department of Chemistry, University of Wisconsin--Madison, 53706, USA.
authors.forename: L
authors.initials: L
authors.lastname: Serfass
authors.forename: H S
authors.initials: HS
authors.lastname: Carr
authors.forename: L M
authors.initials: LM
authors.lastname: Aschenbrenner
authors.forename: J N
authors.initials: JN
authors.lastname: Burstyn
mesh_terms: D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D002413:Cations, Divalent / Q000378:metabolism / Q000494:pharmacology; D006152:Cyclic GMP / Q000378:metabolism*; D015536:Down-Regulation; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008956:Models, Chemical; D009569:Nitric Oxide / Q000378:metabolism; D012995:Solubility
chemical_list: D002413:Cations, Divalent; D009569:Nitric Oxide; D006162:Guanylate Cyclase; D006152:Cyclic GMP; D008274:Magnesium; D002118:Calcium