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• The nature of the primary photochemical events in rhodopsin ...

  The nature of the primary photochemical events in rhodopsin and isorhodopsin Biophysical Journal, Volume 53, Issue 3, 1 March 1988, Pages 367-385 R.R. Birge, C.M. Einterz, H.M. Knapp and L.P. Murray Abstract The nature of the primary photochemical events in rhodopsin and isorhodopsin is studied by using low temperature actinometry, low temperature absorption spectroscopy, and intermediate neglect of differential overlap including partial single and double configuration interaction (INDO-PSDCI) molecular orbital theory. The principal goal is a better understanding of how the protein binding site influences the energetic, photochemical, and spectroscopic properties of the bound chromophore. Absolute quantum yields for the isorhodopsin (I) to bathorhodopsin (B) phototransformation are assigned at 77 K by using the rhodopsin (R) to bathorhodopsin phototransformation as an internal standard (phi R----B = 0.67). In contrast to rhodopsin photochemistry, isorhodopsin displays a wavelength dependent quantum yield for photochemical generation of bathorhodopsin at 77 K. Measurements at seven wavelengths yielded values ranging from a low of 0.089 +/- 0.021 at 565 nm to a high of 0.168 +/- 0.012 at 440 nm. An analysis of these data based on a variety of kinetic models suggests that the I----B phototransformation encounters a small activation barrier (approximately 0.2 kcal mol-1) associated with the 9-cis----9-trans excited-state torsional-potential surface. The 9-cis retinal chromophore in solution (EPA, 77 K) has the smallest oscillator strength relative to the other isomers: 1.17 (all-trans), 0.98 (9-cis), 1.04 (11-cis), and 1.06 (13-cis). The effect of conformation is quite different for the opsin-bound chromophores. The oscillator strength of the lambda max absorption band of I is observed to be anomalously large (1.11) relative to the lambda max absorption bands of R (0.98) and B (1.07). The wavelength-dependent photoisomerization quantum yields and the anomalous oscillator strength associated with isorhodopsin provide important information on the nature of the opsin binding site. Various models of the binding site were tested by using INDO-PSDCI molecular orbital theory to predict the oscillator strengths of R, B, and I and to calculate the barriers and energy storage associated with the photochemistry of R and I for each model. Our experimental and theoretical investigation leads to the following conclusions: (a) The counterion (abbreviated as CTN) is not intimately associated with the imine proton in R, B, or I. The counterion lies underneath the plane of the chromophore in R and I, and the primary chromophore-counterion electrostatic interactions involve C15-CTN and C13-CTN. These interactions are responsible for the anomalous oscillator strength of I relative to R and B.(ABSTRACT TRUNCATED AT 400 WORDS) Abstract | PDF (2207 kb)

• Photochemistry of rhodopsin and isorhodopsin investigated on...

  Photochemistry of rhodopsin and isorhodopsin investigated on a picosecond time scale Biophysical Journal, Volume 27, Issue 1, 1 July 1979, Pages 105-115 T.G. Monger, R.R. Alfano and R.H. Callender Abstract Bovine rhodopsin and isorhodopsin were excited with a single 530-nm, 7-ps light pulse emitted by a mode-locked Nd 3+ glass laser at room temperature. Within 3 ps of excitation, absorbance changes due to formation of bathorhodopsin were observed. The difference spectra generated during and 100 ps after pulse excitation are presented. The data show that bathorhodopsin formation is completed within 3 ps for both the primary pigments and suggest that a single common bathorhodopsin is photochemically formed from both primary pigments. Our findings provide additional support for the cis-trans isomerization model of the primary event in vision. Additional absorption transients that were observed near 670 and 460 nm are discussed. Abstract | PDF (875 kb)

• Photolysis intermediates of the artificial visual pigment ci...

  Photolysis intermediates of the artificial visual pigment cis-5,6-dihydro-isorhodopsin Biophysical Journal, Volume 55, Issue 2, 1 February 1989, Pages 233-241 A. Albeck, N. Friedman, M. Ottolenghi, M. Sheves, C.M. Einterz, S.J. Hug, J.W. Lewis and D.S. Kliger Abstract The photolysis intermediates of an artificial bovine rhodopsin pigment, cis-5,6-dihydro-isorhodopsin (cis-5,6,-diH-ISORHO, lambda max 461 nm), which contains a cis-5,6-dihydro-9-cis-retinal chromophore, are investigated by room temperature, nanosecond laser photolysis, and low temperature irradiation studies. The observations are discussed both in terms of low temperature experiments of Yoshizawa and co-workers on trans-5,6-diH-ISORHO (Yoshizawa, T., Y. Shichida, and S. Matuoka. 1984. Vision Res. 24: 1455–1463), and in relation to the photolysis intermediates of native bovine rhodopsin (RHO). It is suggested that in 5,6-diH-ISORHO, a primary bathorhodopsin intermediate analogous to the bathorhodopsin intermediate (BATHO) of the native pigment, rapidly converts to a blue-shifted intermediate (BSI, lambda max 430 nm) which is not observed after photolysis of native rhodopsin. The analogs from lumirhodopsin (LUMI) to meta-II rhodopsin (META-II) are generated subsequent to BSI, similar to their generation from BATHO in the native pigment. It is proposed that the retinal chromophore in the bathorhodopsin stage of 5,6-diH-ISORHO is relieved of strain induced by the primary cis to trans isomerization by undergoing a geometrical rearrangement of the retinal. Such a rearrangement, which leads to BSI, would not take place so rapidly in the native pigment due to ring-protein interactions. In the native pigment, the strain in BATHO would be relieved only on a longer time scale, via a process with a rate determined by protein relaxation. Abstract | PDF (1065 kb)

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Abstract

Bathorhodopsins were prepared by partially (10--15%) photoconverting bovine rhodopsin (11-cis chromophore) or isorhodopsin I (9-cis chromophore) at 77 degrees K; care was taken to avoid establishing photostationary states. The absorption spectra calculated for the bathorhodopsins derived from the two parent pigments are identical in their lambda max 'S, bandwidths, and extinction coefficients. This result provides further support for the hypothesis that bathorhodopsin is a common intermediate between an 11-cis pigment (rhodopsin) and a 9-cis one (isorhodopsin I) and thus probably has an all-trans chromophore. This in turn is strong evidence for the cis-trans isomerization model of the primary event in vision. The spectrum of the bathoproduct of isorhodopsin II (9,13-dicis chromophore) is different from the other pigments' bathoproducts.