Topography of rhodopsin in rod outer segment disk membranes. Photochemical labeling with N-(4-azido-2-nitrophenyl)-2-aminoethanesulfonate.
Rod cell disk membranes have been photochemically reacted with the water-soluble, membrane-impermeable nitrene precursor N-(4-azido-2-nitrophenyl)-2-aminoethane-sulfonate [NAP-taurine, NAPT]. Rhodopsin, minor membrane proteins, and lipids all incorporate the (nitrophenyl)[35S]taurine (NPT) label. We also find that rhodopsin may be labeled in the dark using prephotolyzed NAPT. This reaction is presumably due to long-lived photoproducts of NAPT. NAPT modifies rhodopsin in the membrane in a selective manner; some cyanogen bromide peptides of NPT-rhodopsin contain appreciable NPT label and some contain essentially none. Precise specific radioactivities could not be determined for the [35S]NPT-peptides since loss of label from some of the peptides was observed during purification procedures. Rhodopsin's carboxyl-terminal cyanogen bromide peptides are well labeled when the protein is modified in disk membranes but the amino-terminal peptide is poorly labeled. When rhodopsin is labeled in reconstituted membranes in which both surfaces of rhodopsin are accessible to reagent, labeling of rhodopsin's amino-terminal peptide is enhanced. These results are consistent with a model in which rhodopsin's carboxyl-terminal region is located at the cytoplasmic (external) surface of disk membranes and its amino terminus is located at the intradiskal membrane surface.
D000345:Affinity Labels; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000032:analysis / Q000648:ultrastructure; D003488:Cyanogen Bromide; D007700:Kinetics; D010446:Peptide Fragments / Q000032:analysis; D010786:Photoreceptor Cells / Q000032:analysis* / Q000648:ultrastructure; D012168:Retinal Pigments / Q000032:analysis*; D012243:Rhodopsin / Q000032:analysis*; D013654:Taurine / Q000031:analogs & derivatives*
D000345:Affinity Labels; D010446:Peptide Fragments; D012168:Retinal Pigments; D013654:Taurine; C010861:NAP taurine; D012243:Rhodopsin; D003488:Cyanogen Bromide