Structural investigations of native and mutant human green cone pigment

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DESCRIPTION (provided by applicant): The green cone pigment absorbs light maximally at a wavelength of 530 nm. Three cone pigments in the retina with distinct light absorbance spectra make trichromatic color vision possible. Despite having unique absorbance spectra, these three pigments utilize the same chemical chromophore, 11-cis-retinal. Absorbance of light isomerizes 11-cis-retinal to all-trans-retinal and activates the pigment protein to propagate the signal down the visual cascade. Understanding how green cone pigment specifically affects the chromophore to maximally absorb 530 nm wavelength light will reveal a basic mechanism of visual function. This will be accomplished by determining a high resolution protein structure by X-ray crystallography together with detailed quantum chemical calculations. The structural conformation and chemical interactions revealed by combining experiments with theory will demonstrate the influence of the protein on the chromophore. With these new insights, two mutations will be introduced and characterized for their influence on the structure of the green cone pigment. The W177R green pigment mutation prevents the protein from folding properly, causing the protein to be retained in the endoplasmic reticulum. This gross structural perturbation will be investigated using the Rosetta protein structure prediction suite. Energetic calculations will reveal key interactions lacking in the W177R mutation that are required for proper protein folding. The A180S mutation does not alter folding of the green cone pigment. To understand the mechanism by which the A180S mutation alters absorbance, we will determine the structure of the mutant pigment experimentally by X-ray crystallography. As with native green cone pigment, a combination of experiment and theory will be used to understand the structural and chemical effects of the A180S mutation. Very few pathogenic mutations have been characterized for cone pigments, a critical step in developing potential therapeutics for deficiencies in human color vision.