Structure of the retinal chromophore in 7,9-dicis-rhodopsin.
Bovine rhodopsin was bleached and regenerated with 7,9-dicis-retinal to form 7,9-dicis-rhodopsin, which was purified on a concanavalin A affinity column. The absorption maximum of the 7,9-dicis pigment is 453 nm, giving an opsin shift of 1600 cm-1 compared to 2500 cm-1 for 11-cis-rhodopsin and 2400 cm-1 for 9-cis-rhodopsin. Rapid-flow resonance Raman spectra have been obtained of 7,9-dicis-rhodopsin in H2O and D2O at room temperature. The shift of the 1654-cm-1 C = N stretch to 1627 cm-1 in D2O demonstrates that the Schiff base nitrogen is protonated. The absence of any shift in the 1201-cm-1 mode, which is assigned as the C14-C15 stretch, or of any other C-C stretching modes in D2O indicates that the Schiff base C = N configuration is trans (anti). Assuming that the cyclohexenyl ring binds with the same orientation in 7,9-dicis-, 9-cis-, and 11-cis-rhodopsins, the presence of two cis bonds requires that the N-H bond of the 7,9-dicis chromophore points in the opposite direction from that in the 9-cis or 11-cis pigment. However, the Schiff base C = NH+ stretching frequency and its D2O shift in 7,9-dicis-rhodopsin are very similar to those in 11-cis- and 9-cis-rhodopsin, indicating that the Schiff base electrostatic/hydrogen-bonding environments are effectively the same. The C = N trans (anti) Schiff base geometry of 7,9-dicis-rhodopsin and the insensitivity of its Schiff base vibrational properties to orientation are rationalized by examining the binding site specificity with molecular modeling.
Loppnow
G R
GR
Department of Chemistry, University of California, Berkeley 94720.
Miley
M E
ME
Mathies
R A
RA
Liu
R S
RS
Kandori
H
H
Shichida
Y
Y
Fukada
Y
Y
Yoshizawa
T
T
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002846:Chromatography, Affinity; D003903:Deuterium; D006860:Hydrogen Bonding; D008958:Models, Molecular; D012160:Retina / Q000737:chemistry*; D012243:Rhodopsin / Q000737:chemistry*; D012374:Rod Cell Outer Segment / Q000737:chemistry; D013059:Spectrum Analysis, Raman; D013696:Temperature; D014867:Water
D014867:Water; D012243:Rhodopsin; D003903:Deuterium