Recent molecular and biochemical studies of the vertebrate visual system have shown that a number of retinal diseases result from mutations in the opsin molecule. Some of these mutations appear to affect either opsin synthesis, insertion into the membrane or degradation, while others affect the visual pigment function in signal transduction. A better correlation between primary structure and resultant opsin function is essential to a better understanding of the genetic lesions in diseased humans, in anticipation of potential gene therapy approaches in the future. The proposed project is designed to identify the molecular basis of wavelength determination in rhodopsin. Rhodopsins of all terrestrial vertebrates absorb at approximately 500 nm. The max is set by the interaction of the chromophore ligand, 11-cis retinal, with the binding pocket of the apoprotein, opsin. Experimental and theoretical studies have resulted in a point-charge model for wavelength modulation, which predicts that negative amino acids in the binding pocket determine the wavelength sensitivity. To test this model, investigators have changed the amino acid residues in the protein's binding pocket by site-specific mutation of the DNA encoding bovine rhodopsin. These data have not yet been successful in generating predictable changes in the wavelength characteristics of the engineered rhodopsin. In this proposal, a novel approach is presented that takes advantage of wavelength shifts that have been selected and fixed through evolutionary pressure. Because of water's characteristic absorption of short and long wavelengths of light, the aquatic environment is increasingly blue with depth. As a result, selection on the wavelength absorption maxima of fishes has shifted them further towards the blue as mean depth occurrence of a fish species increases. To assess how wavelength maxima have been modified in evolutionary """"""""experiments"""""""", fish species have been chosen that have significantly distinct absorption maxima, but that are closely related, so that neutral substitutions are minimized. The rhodopsin genes of eight species of Hawaiian squirrelfishes, whose maxima range from 481 nm to 502 nm, will be sequenced to determine what amino acid residues are responsible for wavelength shifts in the visual pigment. These findings will then be confirmed through the use of site-specific mutation, and spectral analyses of the mutant protein products.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
1R01EY009386-01A1
Application #
3266804
Study Section
Visual Sciences C Study Section (VISC)
Project Start
1992-08-01
Project End
1995-07-31
Budget Start
1992-08-01
Budget End
1993-07-31
Support Year
1
Fiscal Year
1992
Total Cost
Indirect Cost
Name
University of Southern California
Department
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Los Angeles
State
CA
Country
United States
Zip Code
90089