title: Mapping sites of interaction between rhodopsin and transducin using rhodopsin antipeptide antibodies.
abstract: Site-directed antipeptide antibodies generated against the predicted cytoplasmic sequences of rhodopsin were used to map the binding domains for transducin, the retinal G-protein, on the photoreceptor. Antibodies against synthetic peptides corresponding to loop 3-4, loop 5-6, and the serine/threonine-rich region of the COOH terminus recognize rhodopsin by immunoblot analysis and also recognize the native protein within the membrane, allowing these probes to be used for functional studies. Rhodopsin reconstituted into phospholipid vesicles binds transducin in the light which significantly reduces the binding of antipeptide antibodies corresponding to loop 3-4 and the COOH terminus of rhodopsin. However, the binding of the antibody raised against a 14-amino-acid peptide corresponding to a sequence within loop 5-6 of rhodopsin was unaffected by the presence of transducin. These results suggest a preferential involvement of regions in or near loop 3-4 and the COOH terminus in the binding of transducin to rhodopsin. In contrast, a significant portion of loop 5-6 does not form a binding domain for the G-protein.
authors.affiliation: Department of Biochemistry, University of Massachusetts Medical Center, Worcester 01655.
authors.forename: E R
authors.initials: ER
authors.lastname: Weiss
authors.forename: D J
authors.initials: DJ
authors.lastname: Kelleher
authors.forename: G L
authors.initials: GL
authors.lastname: Johnson
mesh_terms: D000818:Animals; D000906:Antibodies; D001665:Binding Sites; D007163:Immunosorbent Techniques; D008565:Membrane Proteins / Q000378:metabolism*; D011817:Rabbits; D012168:Retinal Pigments / Q000378:metabolism*; D012243:Rhodopsin / Q000378:metabolism*; D015293:Transducin
chemical_list: D000906:Antibodies; D008565:Membrane Proteins; D012168:Retinal Pigments; D012243:Rhodopsin; D015293:Transducin