I. Structural studies on rhodopsin by photoaffinity labeling. II. Bioorganic studies on a rhodopsin analog from a retinal containing a 9-membered ring in the side chain

Thesis/Dissertation · Sat Dec 31 23:00:00 EST 1988

OSTI ID:6795065

Sastry, L

Photoaffinity labeling of rhodopsin was carried out to identify the amino acids in the binding sites of the retinal and thereby arrive at a possible helical arrangement of the protein. A retinal analog, with a radioactive photolabel (diazoacetate) at position 3 was used for these studies. {sup 14}C label 3S-diazoacetoxy-9-cis-retinal bound to bovine opsin and regenerated a chromophore with {lambda}{sub max} at 465 nm. Photolysis of the complex at 254 nm resulted in covalent crosslinking of the retinal analog to the protein in 18-20% yield. Proteolytic cleavage (V8 protease) of the crosslinked protein and determination of the distribution of radioactivity indicated that both fragments V8-L (Met{sub 1}-Glu{sub 239}) and V8-S (Ser{sub 240}-Clu{sub 341}) were labeled. Further cleavage of labeled V8-S with CNBr (in HCOOH) showed that the major crosslinking sites were contained in a 51 residue peptide in helix 6, CNBr c+d (Val{sub 258}-Met{sub 308}). An attempt was made to construct a chemical model for bathorhodopsin, the primary photochemical intermediate in the bleaching sequence of rhodopsin.

• Research Organization: Columbia Univ., New York, NY (USA)
• OSTI ID: 6795065
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMALS
BODY
BODY AREAS
CARBON 14 COMPOUNDS
CATTLE
CHEMICAL REACTIONS
CROSS-LINKING
DECOMPOSITION
DOMESTIC ANIMALS
EYES
FACE
HEAD
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
LABELLING
MAMMALS
MOLECULAR STRUCTURE
ORGANIC COMPOUNDS
ORGANS
PHOTOCHEMICAL REACTIONS
PHOTOLYSIS
PIGMENTS
POLYMERIZATION
PROTEINS
RETINA
RHODOPSIN
RUMINANTS
SENSE ORGANS
TRACER TECHNIQUES
VERTEBRATES