Jpn. J. Pharmacol. 69, 239-250 (1995)

Mechanism and Properties of Inhibition of Purified Rat Brain Adenylate Cyclase by G Protein betagamma-Subunits

Keiichi Enomoto, Masako Takano, Shunji Ariyoshi and Takeo Asakawa (*)

Department of Pharmacology, Saga Medical School, Nabeshima, Saga 849, Japan
(*) To whom correspondence should be addressed.

Abstract: The mode of the inhibition of purified rat brain adenylate cyclase by the betagamma-subunits of G protein (betagamma) was studied. These subunits inhibited the catalytic activity of the cyclase with the maximal inhibition of 85% and the half-maximal inhibition at about 0.7 nM betagamma. The complex of betagamma and adenylate cyclase isolated by density gradient centrifugation contained 1.8-2.0 mol beagamma per mol of the cyclase when betagamma was assayed by immunoblotting and by its inhibitory activity on adenylate cyclase. However,the betagamma concentration-inhibition curves suggest that one of the two betagamma molecules bound may be essential for the inhibition. The role for the second betagamma molecule is unknown. As a tentative estimate, 70%, of the adenylate cyclase activity remained inhibited by betagamma when the complex was isolated. The inhibition was not dependent on Galphas or calmodulin. Although purified adenylate cyclase contained a protein (0.06-0.08 mol/mol of adenylate cyclase) that reacted with anti-Galphas antibody, this protein was not liberated from the cyclase when it formed a complex with betagamma. In addition, guanine nucleotide analogs little affected the cyclase activity or the inhibition by betagamma. The inhibition by betagamma was reversed by the dilution of the complex, and the following re-addition of betagamma suppressed the enzyme activity to about 15% of the initial activity again. These findings provide strong evidence that betagamma inhibits adenylate cyclase directly and reversibly through the formation of the complex.

Keywords: Adenylate cyclase, Enzyme inhibition, G protein, betagamma-Subunits, Brain (rat)

Copyright© The Japanese Pharmacological Society 1995

[Back to TOC]