Activation of transducin by a Xenopus short wavelength visual pigment.
Phototransduction in cones differs significantly from that in rods in sensitivity, kinetics, and recovery following exposure to light. The contribution that the visual pigment makes in determining the cone response was investigated biochemically by expressing a Xenopus violet cone opsin (VCOP) cDNA in COS1 cells and assaying the light-dependent activation of transducin. Light-exposed VCOP stimulated [35S]guanosine 5'-(gamma-thio)triphosphate nucleotide exchange on bovine rod transducin in a time-dependent manner with a half-time for activation of 0.75 min, similar to that of bovine rhodopsin. In exhaustive binding assays, VCOP and rhodopsin activity showed similar concentration dependence with half-maximal activation occurring at 0.02 mol of pigment/mol of transducin. Although VCOP was able to activate as many as 12 transducins per photoisomerization, rhodopsin catalyzed significantly more. When assays were performed with lambda > 420 nm illumination, VCOP exhibited rapid regeneration and high affinity for the photoregenerated 11-cis-retinal. Recycling of the chromophore and reactivation of the pigment resulted in multiple activations of transducin, whereas a maximum of 1 transducin per VCOP was activated under brief illumination. The decay of the active species formed following photobleaching was complete in <5 min, approximately 10-fold faster than that of rhodopsin. In vitro, VCOP activated rod transducin with kinetics and affinity similar to those of rhodopsin, but the active conformation decayed more rapidly and the apoprotein regenerated more efficiently with VCOP than with rhodopsin. These properties of the violet pigment may account for much of the difference in response kinetics between rods and cones.
Starace
D M
DM
Department of Biochemistry and Molecular Biology, State University of New York Health Science Center, Syracuse 13210, USA.
Knox
B E
BE
D000426:Alcohol Dehydrogenase / Q000378:metabolism; D000818:Animals; D002417:Cattle; D005960:Glucosides; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D008027:Light; D008099:Liver / Q000201:enzymology; D009243:NAD / Q000378:metabolism; D017299:Rod Opsins / Q000378:metabolism*; D013054:Spectrophotometry, Atomic; D015293:Transducin / Q000378:metabolism*; D014981:Xenopus
D005960:Glucosides; D017299:Rod Opsins; D009243:NAD; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C040358:dodecyl maltoside; D000426:Alcohol Dehydrogenase; D015293:Transducin