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J Biol Chem. 1997 Aug 29;272(35):22265-71.

Interaction of Gsalpha with the cytosolic domains of mammalian adenylyl cyclase.

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  • 1Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas 75235-9041, USA.

Abstract

Forskolin- and Gsalpha-stimulated adenylyl cyclase activity is observed after mixture of two independently-synthesized approximately 25-kDa cytosolic fragments derived from mammalian adenylyl cyclases (native Mr approximately 120,000). The C1a domain from type V adenylyl cyclase (VC1) and the C2 domain from type II adenylyl cyclase (IIC2) can both be expressed in large quantities and purified to homogeneity. When mixed, their maximally stimulated specific activity, 150 micromol/min/mg protein, substantially exceeds values observed previously with the intact enzyme. A soluble, high-affinity complex containing one molecule each of VC1, IIC2, and guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS)-Gsalpha is responsible for the observed enzymatic activity and can be isolated. In addition, GTPgammaS-Gsalpha interacts with homodimers of IIC2 to form a heterodimeric complex (one molecule each of Gsalpha and IIC2) but not detectably with homodimers of VC1. Nevertheless, Gsalpha can be cross-linked to VC1 in the activated heterotrimeric complex of VC1, IIC2, and Gsalpha, indicating its proximity to both components of the enzyme that are required for efficient catalysis. These results and those in the accompanying report (Dessauer, C. W., Scully, T. T., and Gilman, A. G. (1997) J. Biol. Chem. 272, 22272-22277) suggest that activators of adenylyl cyclase facilitate formation of a single, high-activity catalytic site at the interface between C1 and C2.

PMID:
9268375
[PubMed - indexed for MEDLINE]
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