Abstract
The biochemical properties of three stimulatory guanine nucleotide-binding protein (G-protein) alpha subunits, the large and small forms of Gs, Gs-l (52 kDa) and Gs-s (45 kDa), and the olfactory specific G-protein, Golf, have been compared. Complementary DNAs (cDNAs) encoding each alpha subunit were independently expressed in a mammalian cell line deficient in endogenous stimulatory G-proteins (S49 cyc-kin-). Gs-l and Gs-s respond similarly to activation by the beta-adrenergic agonist isoproterenol (EC50 = 80 and 60 nM, respectively) and the receptor-independent G-protein activators guanosine 5-O-3-(thio)triphosphate) (GTP gamma S) and AlF-4. The ability of Golf to interact with the beta-adrenergic receptor was also examined. Surprisingly, Golf interacts with beta-adrenergic receptors and is activated by isoproterenol (EC50 = 120 nM). All three G-proteins respond similarly to treatment with different alpha, beta, and gamma thiophosphoryl analogs of GTP. Specifically, (R)-GTP alpha S and GTP gamma S activate each G-protein, whereas (S)-GTP alpha S and (R)- or (S)-GTP beta S are inactive. In addition, similar to Gs alpha, Golf alpha is covalently modified and constitutively activated by cholera toxin. These studies demonstrate that all three stimulatory G-proteins are functionally and structurally similar, however, subtle differences between Golf and the two forms of Gs appear to modulate their interactions with receptors.
Highlights
The ability of GoIf to interact with the fi-adrenergic receptor was examined
The expression of G,f in S49 cyc-kin- cells allowed us to examine whether G,rcan interact with the @-adrenergic receptor
GOff-containing membranes respond to treatment with isoproterenol (Fig. 1). This indicates that GoIf can couple to @-adrenergic receptors and functionally reconstitute the @-aclrenergic cascade
Summary
The biochemical properties of three stimulatory guanine nucleotide-binding protein (G-protein) a subunits, the large and small forms of Gs, Gs-i (52 kDa) and Gs+. DNAs (cDNAs) encoding each a subunit were independently expressed in a mammalian cell line deficient in endogenous stimulatory G-proteins (S49 cyc-kin-). G%-t and Gs+ respond to activation by the j3-adrenergic agonist isoproterenol (EC&, = 80 and 60 nM, respectively) and the receptor-independent. The ability of GoIf to interact with the fi-adrenergic receptor was examined. All three G-proteins respond to treatment with different a, @, and 7 thiophosphoryl analogs of GTP. (IQ-GTPaS and GTPTS activate each G-protein, whereas
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