| Research Abstract |
Bacteriorhodopsin is a protein which carries our unidirectional proton transport by use of light energy. We have analyzed structural features in this process by Fourier transform infrared spectroscopy, which is pertinent to obtain the informations on the chemical bonds relevant for the function. By locatiog them to the specific positions in the protein by use of site-specific mutant proteins and isotope substitutions, we can reveal the mechanism for the functional expression. Among the photointermediates, both the L intermediate for the proton transfer from the schiff base to Asp85, and the M intermediate for the proton transfer from Asp96 to the Schiff base are important for the proton pumping. It is our prominent contribution to reveal the involvement of the water molecules in these processes. we also revealed the residue responsible for the proton release in the L-to-M conversion as Glu204, which faces to the extracellular medium. By use of mutant proteins with long-lived O intermediate, we showed that the canges in the protein can occur earlier than the changes in the chromophore. A mutant protein, in which Asp85 is replaced by Thr, acquires chloride pumping activity in place of proton pumping activity. On this basis, we revealed transport mechanism of chloride in halorhodopsin. Visual pigment, rhodopsin, has a seven-helical bundle. Similar idea for bacteriorhodopsin was applied to reveal the structure and role of water molecule in the active center of rhodosin. The involvement of water molecules and peptide carbonyls in the process for the activation metarhodopsin II with the G-protein were also elucidated.
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