Calcium regulation of guanine nucleotide activation of hepatic adenylate cyclase.
Pretreatment of isolated rat liver plasma membranes by washing with NaHCO3 buffer or by exposure to the chelator ethyleneglycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) with or without the ionophore A23187, produced a decrease in the sensitivity of adenylate cyclase (ATP pyrophosphate-lyase (cyclizing) EC 4.6.1.1) to subsequent stimulation by NaF or guanosine 5'-(beta-gamma-imino)triphosphate (GPP(NH)P). Sensitivity to activation by the nucleotide could be restored by addition of the lyophilized and ashed wash or by addition of Ca2+, Mg2+ or Mn2+. The factor extracted from the membranes by these various treatments which was responsible for loss of stimulation was identified as Ca2+. Determination of the metal ion content of isolated membranes by atomic absorption spectrometry indicated that Ca2+ was the only divalent cation present in sufficient concentration to support persistent activation by either NaF or GPP(NH)P. Pretreatment of liver plasma membranes with trifluoperazine, which inhibits the action of Ca2+-dependent regulator protein in other enzyme systems, reduced GPP(NH)P activation of adenylate cyclase and caused marked depletion of membrane Ca2+. The effects of low concentrations (less than 100 microM) of the phenothiazine could be reversed totally by Ca2+ and partly by regulator protein. At higher concentrations of trifluoperazine, slight restoration of enzyme activation was seen with either agent. The hypothesis is presented that Ca2+ interacts with the nucleotide (GTP or GDP) regulatory site(s) of the adenylate cyclase. This interaction may be regulator-protein-dependent and may be important in determining the sensitivity of the enzyme to nucleotide activation in vivo.
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D004533:Egtazic Acid; D004789:Enzyme Activation; D006150:Guanine Nucleotides / Q000494:pharmacology*; D008099:Liver / Q000201:enzymology*; D008274:Magnesium / Q000378:metabolism; D008297:Male; D008345:Manganese / Q000378:metabolism; D051381:Rats; D014268:Trifluoperazine / Q000494:pharmacology
D006150:Guanine Nucleotides; D014268:Trifluoperazine; D000001:Calcimycin; D008345:Manganese; D004533:Egtazic Acid; D000262:Adenylyl Cyclases; D008274:Magnesium; D002118:Calcium