title: Structural changes in bacteriorhodopsin following retinal photoisomerization from the 13-cis form.
abstract: Bacteriorhodopsin (BR), a light-driven proton pump in Halobacterium salinarum, accommodates two resting forms of the retinylidene chromophore, the all-trans form (AT-BR) and the 13-cis,15-syn form (13C-BR). Both isomers are present in thermal equilibrium in the dark, but only the all-trans form has proton-pump activity. In this study, we applied low-temperature Fourier-transform infrared (FTIR) spectroscopy to 13C-BR at 77 K and compared the local structure around the chromophore before and after photoisomerization with that in AT-BR. Strong hydrogen-out-of-plane (HOOP) vibrations were observed at 964 and 958 cm(-)(1) for the K state of 13C-BR (13C-BR(K)) versus a vibration at 957 cm(-)(1) for the K state of AT-BR (AT-BR(K)). In AT-BR(K), but not in 13C-BR(K), the HOOP modes exhibit isotope shifts upon deuteration of the retinylidene at C15 and at the Schiff base nitrogen. Whereas the HOOP modes of AT-BR(K) were significantly affected by the mutation of Thr89, this was not the case for the HOOP modes of 13C-BR(K). These observations imply that, while the chromophore distortion is localized near the Schiff base in AT-BR(K), it is located elsewhere in 13C-BR(K). By use of [zeta-(15)N]lysine-labeled BR, we identified the N-D stretching vibrations of the 13C-BR Schiff base (in D(2)O) at 2173 and 2056 cm(-)(1), close in frequency to those of AT-BR. These frequencies indicate strong hydrogen bonding of the Schiff base in 13C-BR, presumably with a water molecule as in AT-BR. In contrast, the N-D stretching vibration appears at 2332 and 2276 cm(-)(1) in 13C-BR(K) versus values of 2495 and 2468 cm(-)(1) for AT-BR(K), suggesting that the rupture of the Schiff base hydrogen bond that occurs in AT-BR(K) does not occur in 13C-BR(K). Rotational motion of the Schiff base upon retinal isomerization is probably smaller in magnitude for 13C-BR than for AT-BR. These differences in the primary step are possibly related to the absence of light-driven proton pumping by 13C-BR.
authors.affiliation: Department of Materials Science and Engineering, Nagoya Institute of Technology, Showa-ku, Nagoya 466-8555, Japan.
authors.forename: Noriko
authors.initials: N
authors.lastname: Mizuide
authors.forename: Mikihiro
authors.initials: M
authors.lastname: Shibata
authors.forename: Noga
authors.initials: N
authors.lastname: Friedman
authors.forename: Mordechai
authors.initials: M
authors.lastname: Sheves
authors.forename: Marina
authors.initials: M
authors.lastname: Belenky
authors.forename: Judith
authors.initials: J
authors.lastname: Herzfeld
authors.forename: Hideki
authors.initials: H
authors.lastname: Kandori
mesh_terms: D001436:Bacteriorhodopsins / Q000737:chemistry*; D003903:Deuterium / Q000737:chemistry; D006859:Hydrogen; D007536:Isomerism; D015394:Molecular Structure; D010777:Photochemistry; D011522:Protons; D012160:Retina / Q000378:metabolism*; D012545:Schiff Bases; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*
chemical_list: D011522:Protons; D012545:Schiff Bases; D001436:Bacteriorhodopsins; D006859:Hydrogen; D003903:Deuterium