Pressure-induced isomerization of retinal on bacteriorhodopsin as disclosed by fast magic angle spinning NMR.
Bacteriorhodopsin (bR) is a retinal protein in purple membrane of Halobacterium salinarum, which functions as a light-driven proton pump. We have detected pressure-induced isomerization of retinal in bR by analyzing 15N cross polarization-magic angle spinning (CP-MAS) NMR spectra of [zeta-15N]Lys-labeled bR. In the 15N-NMR spectra, both all-trans and 13-cis retinal configurations have been observed in the Lys N(zeta) in protonated Schiff base at 148.0 and 155.0 ppm, respectively, at the MAS frequency of 4 kHz in the dark. When the MAS frequency was increased up to 12 kHz corresponding to the sample pressure of 63 bar, the 15N-NMR signals of [zeta-15N]Lys in Schiff base of retinal were broadened. On the other hand, other [zeta-15N]Lys did not show broadening. Subsequently, the increased signal intensity of [zeta-15N]Lys in Schiff base of 13-cis retinal at 155.0 ppm was observed when the MAS frequency was decreased from 12 to 4 kHz. These results showed that the equilibrium constant of [all-trans-bR]/[13-cis-bR] in retinal decreased by the pressure of 63 bar. It was also revealed that the structural changes induced by the pressure occurred in the vicinity of retinal. Therefore, microscopically, hydrogen-bond network around retinal would be disrupted or distorted by a constantly applied pressure. It is, therefore, clearly demonstrated that increased pressure induced by fast MAS frequencies generated isomerization of retinal from all-trans to 13-cis state in the membrane protein bR.
Kawamura
Izuru
I
Graduate School of Engineering, Yokohama National University, Hodogaya-ku, Yokohama, Japan.
Degawa
Yoshiaki
Y
Yamaguchi
Satoru
S
Nishimura
Katsuyuki
K
Tuzi
Satoru
S
Saitô
Hazime
H
Naito
Akira
A
D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000528:radiation effects; D019617:Halobacterium salinarum / Q000737:chemistry / Q000528:radiation effects; D006860:Hydrogen Bonding; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D010777:Photochemistry; D011312:Pressure; D012172:Retinaldehyde / Q000737:chemistry; D012545:Schiff Bases / Q000737:chemistry; D013237:Stereoisomerism
D012545:Schiff Bases; D001436:Bacteriorhodopsins; D012172:Retinaldehyde