PHOTOPHYSICS OF RHODOPSIN AND BACTERIORHODOPSIN

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Rhodopsin, the protein responsible for converting light into an optic nerve impulse, and bacteriorhodopsin, the light transducing protein of the purple membrane of Halobacterium halobium, have significantly different biological roles. Nevertheless, natural selection has converged on very similar designs for both proteins, and these similarities have prompted comparative experimental and theoretical studies. The lack of high resolution X-ray structural data has precluded precise assignment of the tertiary structure of either protein, and hence our knowledge of the binding sites and the primary events is based on indirect analyses of chemical and spectroscopic data. Despite a number of important advances during the past decade much remains to be understood and a number of interesting experimental paradoxes remain unexplained. The nature of the chromophore binding sites and the nature of the primary photochemical events of rhodopsin and bacteriorhodopsin will be studied by using spectroscopic and theoretical techniques. The goals are to understand the photophysical properties of the bound chromophores and how the proteins mediate the photochemical properties of the bound chromophores. The principal experimental methods to be used in these studies include two-photon spectroscopy, Stark effect spectroscopy, Fourier transform near infra-red spectroscopy, microwave spectroscopy, pulsed laser photocalorimetry, and time-resolved photovoltaic spectroscopy. The theoretical methods include semiempirical molecular orbital theory and molecular dynamics theory. In addition, we will seek to answer the following specific questions: (1) To what extent does water participate in mediating the photochemical properties of the bound chromophores in both proteins? (2) What is the origin of the fast photoelectric signal that is generated during the primary photochemical events in rhodopsin and bacteriorhodopsin? (3) Can we observe and use protein-chromophore charge transfer bands to help analyze the nature of the protein chromophore interactions within the protein binding site? (4) What is the origin of the strong microwave absorptivity of bacteriorhodopsin and the origin of the M - bR microwave difference spectrum? (5) What is the nature of energy storage in the primary events of rhodopsin and bacteriorhodopsin? (6) Are there charged amino acids near the ring of the chromophore in bacteriorhodopsin? Furthermore, we will seek to develop a new method of collecting two-photon spectra of photochemically labile biological molecules based on Fourier transform optical techniques.