Adenylate kinase increases adenylate cyclase activity in membranes from rat lung.
The adenylate cyclase activity of membranes prepared from rat lung, measured under standard assay conditions, was markedly increased by the presence of a crude supernatant fraction prepared from rat lung, liver, or brain. This was not due to an increase in the initial rate of cyclic AMP (cAMP) synthesis, but to the maintenance of a constant rate of cAMP synthesis for periods of at least 10 min. After incubating lung membranes in the cyclase reaction mixture until cAMP synthesis had virtually ceased (10 min), the addition of alpha-(32P)-ATP caused a marked increase in the activity of the enzyme. This was the only component of the original reaction mixture that supported re-initiation of cAMP synthesis. Re-initiation also occurred when supernatant was added. This implies that substrate depletion occurs in the presence of membranes and that lung supernatant can catalyze rapid regeneration of substrate. Chromatographic analysis confirmed that ATP was rapidly hydrolyzed to AMP in the presence of the membranes, that this rapid destruction of ATP did not occur when supernatant was present, and that ATP was resynthesized from AMP when supernatant was added to a reaction mixture in which most of the ATP initially present had been destroyed. The effects of supernatant were mimicked by commercially available adenylate kinase. Addition of adenylate kinase did not affect adenylate cyclase activity measured in membranes prepared from brain, heart, or kidney, suggesting that lung membranes may contain more nucleotide pyrophosphatase and/or less endogenous adenylate kinase activity. Studies of soluble factors that affect adenylate cyclase must carefully control for differential substrate depletion in the presence and absence of tissue extracts.
D000263:Adenylate Kinase / Q000378:metabolism*; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology; D002462:Cell Membrane / Q000201:enzymology; D066298:In Vitro Techniques; D008099:Liver / Q000201:enzymology; D008168:Lung / Q000201:enzymology*; D008297:Male; D009711:Nucleotides / Q000378:metabolism; D010761:Phosphorus Radioisotopes; D010770:Phosphotransferases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013997:Time Factors
D009711:Nucleotides; D010761:Phosphorus Radioisotopes; D010770:Phosphotransferases; D000263:Adenylate Kinase; D000262:Adenylyl Cyclases