title: Crystal structures of different substrates of bacteriorhodopsin's M intermediate at various pH levels.
abstract: The hexagonal P622 crystal of bacteriorhodopsin, which is made up of stacked membranes, is stable provided that the precipitant concentration in the soaking solution is higher than a critical value (i.e., 1.5 M ammonium sulfate). Diffraction data showed that the crystal lattice shrank linearly with increasing precipitant concentration, due primarily to narrowing of intermembrane spaces. Although the crystal shrinkage did not affect the rate of formation of the photoreaction M intermediate, its lifetime increased exponentially with the precipitant concentration. It was suggested that the energetic barrier of the M-to-N transition becomes higher when the motional freedom of the EF loop is reduced by crystal lattice force. As a result of this property, the M state accumulated predominantly when the crystal that was soaked at a high precipitant concentration was illuminated at room temperature. Structural data obtained at various pH levels showed that the overall structure of M is not strongly dependent on pH, except that Glu194 and Glu204 in the proton release complex are more separated at pH 7 than at pH 4.4. This result suggests that light-induced disruption of the paired structure of Glu194 and Glu204 is incomplete when external pH is lower than the pK(a) value of the proton release group in the M state.
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University, Nagoya, Japan.
authors.forename: Masataka
authors.initials: M
authors.lastname: Yamamoto
authors.forename: Naoki
authors.initials: N
authors.lastname: Hayakawa
authors.forename: Midori
authors.initials: M
authors.lastname: Murakami
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Kouyama
mesh_terms: D000042:Absorption / Q000187:drug effects / Q000528:radiation effects; D000645:Ammonium Sulfate / Q000737:chemistry; D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000378:metabolism; D020134:Catalytic Domain; D011232:Chemical Precipitation; D018360:Crystallography, X-Ray; D019617:Halobacterium salinarum / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration / Q000187:drug effects / Q000528:radiation effects; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D009997:Osmotic Pressure / Q000187:drug effects / Q000528:radiation effects; D011487:Protein Conformation / Q000187:drug effects / Q000528:radiation effects; D011522:Protons; D013057:Spectrum Analysis; D013379:Substrate Specificity / Q000187:drug effects / Q000528:radiation effects; D014965:X-Rays
chemical_list: D011522:Protons; D001436:Bacteriorhodopsins; D000645:Ammonium Sulfate