Dynamic aspect of bacteriorhodopsin as a typical membrane protein as revealed by site-directed solid-state 13C NMR

Saitô, Hazime; Yamaguchi, Satoru; Okuda, Hideyasu; Shiraishi, Aya; Tuzi, Satoru

doi:10.1016/j.ssnmr.2003.06.001

Cited by 10 publications

(6 citation statements)

References 36 publications

(66 reference statements)

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“…During the last few years an expanded series of papers was published [110][111][112][113][114][115][116][117][118][119][120] in which the authors studied dynamic features of bacteriorhodopsin and other membrane proteins by means of site-directed 13 C solid-state NMR spectroscopy. The term 'site directed' means that in the proteins only one carbon of only one type of aminoacid (in most cases, Val and Ala) was labeled.…”

Section: Lineshape and Relaxation Experiments In Isotopically Enrichementioning

confidence: 99%

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Solid-state NMR and protein dynamics

Krushelnitsky

Reichert²

2005

Progress in Nuclear Magnetic Resonance Spectroscopy

102

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Section: Lineshape and Relaxation Experiments In Isotopically Enrichementioning

confidence: 99%

“…However, measuring spectra at many different conditions (labelling patterns, pH, temperatures, etc.) enabled them to obtain a map of mobile regions [115] and to relate the internal dynamics to proton uptake and transport [120] performed by bacteriorhodopsin.…”

Section: Lineshape and Relaxation Experiments In Isotopically Enrichementioning

confidence: 99%

Solid-state NMR and protein dynamics

Krushelnitsky

Reichert²

2005

Progress in Nuclear Magnetic Resonance Spectroscopy

102

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“…We have observed that local conformation and dynamics by 13 C‐NMR study of site specifically isotopic‐labeled bR (15–17). In particular, it turned out that 13 C‐NMR peaks were well resolved for fully hydrated [3‐ 13 C]Ala‐, [1‐ 13 C]Val‐labeled bR, depending upon their local dynamics and conformations of 29 Ala or 21 Val residues, respectively (18,19).…”

Section: Introductionmentioning

confidence: 99%

Pressure-induced Isomerization of Retinal on Bacteriorhodopsin as Disclosed by Fast Magic Angle Spinning NMR†

Kawamura

Degawa

Yamaguchi

et al. 2007

Photochemistry and Photobiology

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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.

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“…For 15 N T 2 values, the N-H dipolar interaction could dominate, as seen when T 2 is gradually increased at low temperature ( Figure 7). Therefore, at 293 K, proton decoupling was not effective because of an interference of the frequency of molecular motion with the frequency of the NH fluctuations (91), resulting in the poor signal-to noise ratio of 15 N CP spectrum, which recovered gradually by cooling, as the molecular motion became slower ( Figure 5). However, here, the dipole-dipole interactions are dependent on the dipolar orientation.…”

Section: Dynamics Of Phospholipids In the Pmmentioning

confidence: 99%

Functionally Relevant Coupled Dynamic Profile of Bacteriorhodopsin and Lipids in Purple Membranes

Kamihira

Watts

2006

Biochemistry

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The dynamics of bacteriorhodopsin (bR) and the lipid headgroups in oriented purple membranes (PMs) was determined at various temperatures and relative humidity (rh) using solid-state NMR spectroscopy. The 31 P NMR spectra of the R-and γ-phosphate groups in methyl phosphatidylglycerophosphate (PGP-Me), which is the major phospholipid in the PM, changed sensitively with hydration levels. Between 253 and 233 K, the signals from a fully hydrated sample became broadened similarly to those of a dry sample at 293 K. The 15 N cross polarization (CP) NMR spectral intensities from [ 15 N]Gly bR incorporated into fully hydrated PMs were suppressed in 15 N CP NMR spectra at 293 K compared with those of dry membranes but gradually recovered at low temperatures or at lower hydration (75%) levels. The suppression of the NMR signals, which is due to interference with proton decoupling frequency (∼45 kHz), coupled with short spin-spin relaxation times (T 2 ) indicates that the loops of bR, in particular, have motional components around this frequency. The motion of the transmembrane R-helices in bR was largely affected by the freezing of excess water at low temperatures. While between 253 and 233 K, where a dynamic phase transition-like change was observed in the 31 P NMR spectra for the phosphate lipid headgroups, the molecular motion of the loops and the C-and N-termini slowed, suggesting lipidloop interactions, although protein-protein interactions between stacks cannot be excluded. The results of T 2 measurements of dry samples, which do not have proton pumping activity, were similar to those for fully hydrated samples below 213 K where the M-intermediates can be trapped. These results suggest that motions in the 10s µs correlation regime may be functionally important for the photocycle of bR, and protein-lipid interactions are motionally coupled in this dynamic regime.Bacteriorhodopsin (bR) 1 is one of the most extensively studied light-driven proton pumps in Haloarchaea. It contains seven transmembrane R-helices surrounding the retinal chromophore and has been the structural paradigm of G-protein-coupled receptors and other membrane proteins (1-5). It forms a protein-lipid complex of defined composition in the purple membrane (PM), where bR is arranged in trimeric units assembled in 2D crystalline patches in a hexagonal lattice. High-resolution X-ray (6-11) and electron (12, 13) crystallography have been used to determine the 3D structure of bR and the structural changes at K, L, M, N, and O photointermediates to understand the unidirectional translocation of proton pumping activity (14-23). Solid-state NMR spectroscopy has been used to provide detailed information about the local structural changes of retinal (24,25) around the key residues involved in proton pumping activity (26,27) and the local mobility of bR (28-30) under physiological conditions. Also, the structure and orientation of some R-helices of membrane-embedded bR have been determined by polarization inversion spin exchange at the magic angle (PISE...

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Dynamic aspect of bacteriorhodopsin as a typical membrane protein as revealed by site-directed solid-state 13C NMR

Cited by 10 publications

References 36 publications

Solid-state NMR and protein dynamics

Solid-state NMR and protein dynamics

Pressure-induced Isomerization of Retinal on Bacteriorhodopsin as Disclosed by Fast Magic Angle Spinning NMR†

Functionally Relevant Coupled Dynamic Profile of Bacteriorhodopsin and Lipids in Purple Membranes

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