Low Temperature FTIR Studies on the Photointermediates L, M, and N of Bacteriorhodopsin

Maeda, Akio; Pfefferlé, J.-M.; Sasaki, Jun; Shichida, Yoshinori; Yoshizawa, Tôru

doi:10.1007/978-3-642-84771-4_20

Cited by 5 publications

(4 citation statements)

References 5 publications

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“…As demonstrated previously for BR, information about structural changes of the protein and chromophore can be obtained by measuring band shifts induced by H/D exchange (Braiman & Rothschild, 1988;Rath et al, 1993). For example, the extent of the H/D-induced downshift of the carboxylic CdO stretch mode of Asp and Glu amino acid residues decreases with stronger hydrogen bonding, reflecting the coupling between the OH in-plane bend and the CdO stretch modes (Maeda et al, 1992). In contrast, ester carbonyl groups of phospholipids such as the L-R-phosphatidylglycerol lipids which were used to reconstitute SR-I in these studies are insensitive to H/D exchange.…”

Section: Resultsmentioning

confidence: 99%

Asp76 Is the Schiff Base Counterion and Proton Acceptor in the Proton-Translocating Form of Sensory Rhodopsin I

et al. 1996

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Both sensory rhodopsin I, a phototaxis receptor, and bacteriorhodopsin, a light-driven proton pump, have homologous residues which have been identified as critical for bacteriorhodopsin functioning. This includes Asp76, which in the case of bacteriorhodopsin (Asp85) functions as both the Schiff base counterion and the proton acceptor. Sensory rhodopsin I exists in a pH dependent equilibrium between two different forms in the absence of its transducer protein HtrI. At pH below 7, it exists primarily in a blue form (lambda max = 587 nm) which functions as a phototaxis signal transducer when complexed to HtrI, while at higher pH, it converts to a purple proton-transporting form similar to bacteriorhodopsin (lambda max = 550 nm). We report ATR-FTIR difference spectra obtained from both low- and high-pH forms of purified sensory rhodopsin I reconstituted into lipid vesicles. The low-pH species has an ethylenic C = C stretch mode at 1520 cm-1 which shifts to 1526 cm-1 in the high-pH form. No frequency shift was found for the mutant D76N, in agreement with visible absorption measurements. Weak negative/positive bands at 1763/1751 cm-1 previously assigned to a perturbation of the C = O stretch mode of Asp76 during S373 formation in the low-pH form are replaced by a single intense positive band near 1749 cm-1 in the high-pH form. These results along with the effects of H/D exchange show that Asp76 is protonated in the signal-transducing form of sensory rhodopsin I and is ionized and functions as the counterion and Schiff base proton acceptor in the proton-transporting high-pH form of sensory rhodopsin I similar to bacteriorhodopsin.

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Section: Resultsmentioning

confidence: 99%

Asp76 Is the Schiff Base Counterion and Proton Acceptor in the Proton-Translocating Form of Sensory Rhodopsin I

et al. 1996

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“…the basis of an H 18 2 O induced frequency shift of V12 cm 31 (Fig. 3) to a weakly hydrogen bonded water molecule which undergoes a further decrease in hydrogen bonding during M formation [36,37].…”

Section: The 3300^3700 CM 31 Regionmentioning

confidence: 99%

Detection of threonine structural changes upon formation of the M-intermediate of bacteriorhodopsin: evidence for assignment to Thr-89

Liu

Lee

Coleman

et al. 1998

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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The behavior of threonine residues in the bacteriorhodopsin (bR) photocycle has been investigated by Fourier transform infrared difference spectroscopy. L-Threonine labeled at the hydroxyl group with 18O (L-[3-(18)O]threonine) was incorporated into bR and the bR-->M FTIR difference spectra measured. Bands are assigned to threonine vibrational modes on the basis of 18O induced isotope frequency shifts and normal mode calculations. In the 3500 cm-1 region, a negative band is assigned to the OH stretch of threonine. In the 1125 cm-1 region, a negative band is assigned to a mixed CH3 rock/CO stretch mode. The frequency of both these bands indicates the presence of at least one hydrogen bonded threonine hydroxyl group in light adapted bR which undergoes a change in structure by formation of the M intermediate. Spectral changes induced by the substitution Thr-89-->Asn but not Thr-46-->Asn or Asp-96-->Asn are consistent with the assignment of these bands to Thr-89. These results along with another related study on the mutant Thr-89-->Asn indicate that the active site of bR includes Thr-89 and that its interaction with the retinylidene Schiff base and Asp-85 may play an important role in regulating the color of bacteriorhodopsin and the transfer of a proton to the Schiff base.

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“…The photoinduced protein conformational change and retinal configurational change of bR during photocycle have been extensively studied by using low-temperature Fourier transform infrared difference spectroscopy and time-resolved Fourier transform infrared spectroscopy in the midinfrared (IR) region (Siebert et al, 1982;Maeda et al, 1992;Weidlich and Siebert, 1993;Wolfgang et al, 1996;Dioumaev and Braiman, 1997;Rö dig et al, 1999). Very recently, the spectrally resolved IR emission of bR has been reported (Terpugov and Degtyareva, 2001): under steady-state photoexcitation, the majority of the observed emission peaks in the mid-IR region have been attributed to the retinal chromophore in the ground state and in the K intermediate.…”

Section: Introductionmentioning

confidence: 99%

Time-Resolved Long-Lived Infrared Emission from Bacteriorhodopsin during its Photocycle

Wang

El‐Sayed

2002

Biophysical Journal

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The infrared emission observed below 2000 cm(-1) upon exciting retinal in bacteriorhodopsin (bR) is found to have a rise time in the submicrosecond time regime and to relax with two exponential components on the submillisecond to millisecond time scale. These time scales, together with the assignment of this emission to hot vibrations from the all-trans retinal (in bR) and the 13-cis retinal (in the K intermediate), support the recent assignment of the J-intermediate as an electronically excited species (Atkinson et al., J. Phys. Chem. A. 104:4130-4139, 2000) rather than a vibrationally hot K intermediate. A discussion of these time scales of the observed infrared emission is given in terms of the competition between radiative and nonradiative relaxation processes of the vibrational states involved.

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Low Temperature FTIR Studies on the Photointermediates L, M, and N of Bacteriorhodopsin

Cited by 5 publications

References 5 publications

Asp76 Is the Schiff Base Counterion and Proton Acceptor in the Proton-Translocating Form of Sensory Rhodopsin I

Asp76 Is the Schiff Base Counterion and Proton Acceptor in the Proton-Translocating Form of Sensory Rhodopsin I

Detection of threonine structural changes upon formation of the M-intermediate of bacteriorhodopsin: evidence for assignment to Thr-89

Time-Resolved Long-Lived Infrared Emission from Bacteriorhodopsin during its Photocycle

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