ACTIVATION OF THE GTP-BINDING PROTEIN GQ BY RHODOPSIN IN SQUID PHOTORECEPTORS

Photoaffinity labelling by a GTP analogue has been used to identify a 42 kDa band as the major Galpha subunit in squid photoreceptor membranes, recently identified by partial sequence analysis to be a member of the Gq sub-group of GTP-binding proteins [Pottinger, Ryba, Keen & Findlay (1991) Biochem. J. 279, 323-326]. Guanine-nucleotide-binding displacement analysis gave a stoichiometry of 1 G-protein per 12.5 rhodopsin molecules, the same as in vertebrate rod photoreceptors. Binding was not detected above background in the dark, but was rapidly activated by light. Unlike vertebrate transducin, this G-protein is very temperature-sensitive. GTP binding is maximal at temperatures less than 10-degrees-C and is much decreased after several minutes above 18-degrees-C. The light-stimulated GTPase rate is maximal around 10-degrees-C, above which the loss of binding sites counteracts the increase in hydrolytic rate per site. Earlier studies described light-sensitive Galpha components of 40 and 45 kDa, by ADP-ribosylation in the presence of cholera and pertussis toxins. These are now shown to be very minor components, as the prolonged treatment at elevated temperature required for ADP-ribosylation is sufficient to inactivate the major Galpha totally. Unlike the minor Galpha components, the 42 kDa Galpha is not inhibited by Ca2+.