Purified guanylate cyclase: characterization, iodination and preparation of monoclonal antibodies.
Guanylate cyclase was purified from the soluble fraction of rat lung using a modification of procedures published previously. The purified enzyme exhibited specific activities, at pH 7.6, of 219-438 nmoles/mg protein/min and 34-60 nmoles/mg protein/min with Mn2+ and Mg2+ as cation cofactors, respectively. The specific activity changed as a function of the protein concentration due to a change in Vmax with no alteration of the Km for GTP. The enzyme migrated as a single band coincident wih guanylate cyclase activity on nondenaturing polyacrylamide and isoelectric focusing gels (isoelectric point = 5.9). Purified guanylate cyclase had an apparent molecular weight of 150,000 daltons as determined by gel filtration chromatography and polyacrylamide gel electrophoresis. Electrophoresis in the presence of sodium dodecyl sulfate revealed a single subunit of 72,000 daltons, suggesting that the enzyme is a dimer of an identical subunit. The purified enzyme could be activated by nitric oxide, indicating that this compound interacts directly with the enzyme.
D000818:Animals; D000906:Antibodies; D055598:Chemical Phenomena; D002621:Chemistry; D002999:Clone Cells / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D006162:Guanylate Cyclase / Q000276:immunology / Q000302:isolation & purification*; D006822:Hybrid Cells / Q000276:immunology; D007700:Kinetics; D008168:Lung / Q000201:enzymology*; D051379:Mice / Q000276:immunology; D009569:Nitric Oxide; D051381:Rats
D000906:Antibodies; D009569:Nitric Oxide; D006162:Guanylate Cyclase