Stabilization and solubilization of bovine corpus-luteum adenylate cyclase. The effects of guanosine triphosphate, guanosine 5'-[beta,gamma-imido]triphosphate, sodium fluoride and Tris/hydrochloric acid concentration on enzyme activity.
1. Adenylate cyclase of the washed 600g sediment of bovine corpus-luteum homogenate is stimulated by p[NH]ppG (guanosine 5'-[beta,gamma-imido]triphosphate), the imido analogue of GTP, and to a lesser extent by GTP itself. Activation by p[NH]ppG is not reversed by extensive washing before assay, but can, however, be reversed by NaF. 2. Both p[NH]ppG and NaF stabilize the enzyme during incubation at 37 degrees C. NaF also causes an irreversible activation, but only of part of the potentially NaF-activatable adenylate cyclase; there are possibly two components of the adenylate cyclase system, which can be distinguished by their response to NaF. 3. Solubilization of the adenylate cyclase activity in the 600g sediment, by using the non-ionic detergent Lubrol-PX, gave variable yields. A relationship between the magnitude of NaF stimulation of the 600g-sediment enzyme and the yield of soluble activity derived from the sediment was recognized. The results suggest that the pre-existing state of the enzyme complex in vivo is reflected by the response in vitro to NaF and may determine the success with which activity can be solubilized. 4. The absolute yields of soluble activity could be increased by p[NH]ppG preactivation of the 600g sediment. During the development of the maximally active state by preincubation with p[NH]ppG the enzyme passes through a stage in which Lubrol solubilization is increased, but the maximally active state is itself less amenable to solubilization. p[NH]ppG activation causes the appearance of NaF-inhibited states, which appear to be preferentially solubilized by Lubrol-PX.
D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D003338:Corpus Luteum / Q000201:enzymology; D003902:Detergents / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D006851:Hydrochloric Acid / Q000494:pharmacology; D008298:Maleates / Q000494:pharmacology; D012969:Sodium Fluoride / Q000494:pharmacology; D012995:Solubility; D014325:Tromethamine / Q000494:pharmacology
D003902:Detergents; D008298:Maleates; D014325:Tromethamine; D006165:Guanylyl Imidodiphosphate; D006160:Guanosine Triphosphate; D012969:Sodium Fluoride; D000262:Adenylyl Cyclases; D006851:Hydrochloric Acid