Interaction of Aspartate-85 with a Water Molecule and the Protonated Schiff Base in the L Intermediate of Bacteriorhodopsin: A Fourier-Transform Infrared Spectroscopic Study

Maeda, Akio; Sasaki, Jun; Yamazaki, Yoichi; Needleman, Richard; Lanyi, Janos Κ.

doi:10.1021/bi00173a013

Cited by 114 publications

(118 citation statements)

References 48 publications

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“…This feature which was initially discovered in studies of L at 170 K and M at 230 K (48) is observed as a depletion band upon formation of L at 298 K, and this feature persists in M and N. This band is also the same one that was recently detected in static difference spectra, obtained using an in situ H 2 16 O/ H 2 18 O exchange technique with BR at room temperature (37). In the H 2 16 O/ H 2 18 O exchange measurements, the band was not observed in D85N (37) and it was also not observed in the light-induced difference spectrum of the D85N mutant at 170 K (25). As discussed previously (37,49) this water vibration is very likely due to the free O-H bond of Wat401 in BR, a water whose other hydrogen interacts with Asp85 (i.e., Asp85-COO − ••H-O-H), as is seen in the crystal structure of BR (50).…”

Section: Water Forms H-bonds With Asp85 In L M and N At 298 Ksupporting

confidence: 83%

Water Structural Changes in the L and M Photocycle Intermediates of Bacteriorhodopsin as Revealed by Time-Resolved Step-Scan Fourier Transform Infrared (FTIR) Spectroscopy

et al. 2007

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In previous FTIR studies of the photocycle intermediates of bacteriorhodopsin at cryogenic temperatures, water molecules were observed in the L intermediate, in the region surrounded by protein residues between the Schiff base and Asp96. In the M intermediate, the water molecules had moved away towards the Phe219-Thr46 region, but in the N intermediate they were again observed near the Schiff base. In order to evaluate the relevance of this scheme at room temperature, time resolved FTIR difference spectra of bacteriorhodopsin, including the water O-H stretching vibration frequency regions, were recorded in the μsec and msec time ranges. Vibrational changes of weakly-H-bonded water molecules were observed in L, M, and N. In each of these intermediates, the depletion of a water O-H stretching vibration at 3645 cm −1 originating from the initial unphotolzyed bacteriorhodopsin was observed as a trough in the difference spectrum. This vibration is due to the dangling O-H group of a water molecule which interacts with Asp85, and its absence in each of these intermediates indicates that there is perturbation of this O-H group. The formation of M is accompanied by the appearance of water O-H stretching vibrations at 3670 and 3657 cm −1 . These bands are due to water molecules present in the region surrounded by Thr46, Asp96 and Phe219. Formation of L at 298 K is accompanied by the perturbations of Asp96 and the Schiff base, though in different ways from what is observed at 170 K. Changes in a broad water vibrational feature, centered around 3610 cm −1 , are kinetically correlated with the L-to-M transition. These results imply that even at room temperature, water molecules interact with Asp96 and the Schiff base in L, though with a less rigid structure than at cryogenic temperatures.Bacteriorhodopsin is a light-driven proton pump. It undergoes a photocycle initiated by the absorption of light by a retinal chromophore which is covalently linked to Lys216 forming a protonated Schiff base. The initial consequence of light absorption is the trans-to-cis isomerization of the C 13 =C 14 bond of the retinal. Proton pumping takes place as the protein undergoes conformational changes from the initial unphotolyzed state with an all-trans retinal chromophore (BR 1 ) through a sequence of intermediates, and back to the BR state. Spectroscopic and kinetic studies, including FTIR studies have distinguished a sequence of + This work was supported by an NIH grant HL 16101 (to RBG).

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Section: Water Forms H-bonds With Asp85 In L M and N At 298 Ksupporting

confidence: 83%

Water Structural Changes in the L and M Photocycle Intermediates of Bacteriorhodopsin as Revealed by Time-Resolved Step-Scan Fourier Transform Infrared (FTIR) Spectroscopy

et al. 2007

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“…Asp-85 is located near the Schiff base and serves as the counterion. The presence of water molecules in this region had been suggested by Fourier transform infrared (FTIR) studies (4,5), and these were later confirmed as W401, W402, and W406 in the high resolution crystal structure of the BR ground state (PDB code 1C3W; see Fig. 1) (6).…”

Section: Bacteriorhodopsin (Br)mentioning

confidence: 81%

Factors That Differentiate the H-bond Strengths of Water Near the Schiff Bases in Bacteriorhodopsin and Anabaena Sensory Rhodopsin

Saito

Kandori

Ishikita

2012

Journal of Biological Chemistry

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Background: Bacteriorhodopsin (BR) functions as a proton pump, whereas Anabaena sensory rhodopsin (ASR) functions as a photosensor. Results: pK a of the conserved Asp residues near the Schiff base significantly differ between BR and ASR. Conclusion:The O-D stretching frequencies for D 2 O are correlated with pK a (Asp). Significance: The presence of a strongly H-bonded water results from the proton-pumping activity in BR.

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“…However, the crystal structure shows a water molecule in the vicinity of the FMN chromophore (7). It has been demonstrated that in bacteriorhodopsin structural water is required to stabilize the protein and participates in intramolecular proton transfer reactions during the proton-pumping photocycle (32,33). This water itself could be either the proton donor/acceptor group, or it could act as a bridge between the cysteine and another remote amino acid side chain that donates/accepts the proton.…”

Section: Protein-chromophore Interactions In the Lov2 Groundmentioning

confidence: 99%

The Photocycle of a Flavin-binding Domain of the Blue Light Photoreceptor Phototropin

Swartz¹,

Corchnoy²,

Christie³

et al. 2001

Journal of Biological Chemistry

518

783

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The plant blue light receptor, phot1, a member of the phototropin family (1), is a plasma membrane-associated flavoprotein that contains two (ϳ110 amino acids) flavinbinding domains, LOV1 and LOV2, within its N terminus and a typical serine-threonine protein kinase domain at its C terminus. The LOV (light, oxygen, and voltage) domains belong to the PAS domain superfamily of sensor proteins. In response to blue light, phototropins undergo autophosphorylation. E. coli-expressed LOV domains bind riboflavin-5-monophosphate, are photochemically active, and have major absorption peaks at 360 and 450 nm, with the 450 nm peak having vibronic structure at 425 and 475 nm. These spectral features correspond to the action spectrum for phototropism in higher plants. Near-UV blue light regulates a variety of different responses in higher plants. These include phototropism, the inhibition of hypocotyl elongation, the expression of various genes, and stomatal opening. Phot1 (nph1), the recently discovered blue light receptor, is a member of the phototropin receptor family (1). Phot1 is a plasma membrane-associated flavoprotein that functions as the primary photoreceptor mediating phototropic plant movement (2-4). Phot1 has two 12.1-kDa flavin-binding domains, LOV1 and LOV2, within its N-terminal region and a typical serinethreonine protein kinase domain at the C-terminal region. Heterologous expression studies have shown that phot1 binds FMN 1 as a chromophore and undergoes autophosphorylation in response to light treatment. It has therefore been proposed that this receptor functions as a light-activated serine/threonine kinase (4). The isolated LOV domains from oat phot1 expressed in Escherichia coli have been shown to undergo a cyclic photoreaction upon the absorption of light; LOV1 recovers with a half-time of 11.5 s, whereas LOV2 recovers with a half-time of 27 s (5). In addition, the quantum efficiencies for photoproduct (adduct) formation for LOV1 and LOV2 are ϳ0.045 and 0.44, respectively (5). The ground forms of the LOV domains have major absorption peaks at 360 and 450 nm with the 450 peak having vibronic structure at 425 and 475 nm. Upon absorption of light, the chromophore bleaches 2 in the 450 nm region generating a species that absorbs maximally at 390 nm. This intermediate has been assigned as a flavin-cysteinyl adduct between the protein and the C(4a) carbon of the FMN chromophore. This adduct breaks down spontaneously, returning the protein to its ground form. A LOV2 mutant (LOV2C39A) in which the cysteine that forms the adduct has been mutated to alanine does not undergo this photoreaction (5).Recently the crystal structure of the LOV2 domain from the fern Adiantum capillus-veneris phy3 (6) was solved to 2.7-Å resolution (7). Phy3 is a chimeric photoreceptor with homology to phytochrome at its N-terminal end and an almost complete phototropin at its C-terminal end. Its LOV2 domain shares a 70% sequence homology to the oat phot1 LOV2 (6). The structure indicates that the FMN molecule is held noncovalently within...

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Interaction of Aspartate-85 with a Water Molecule and the Protonated Schiff Base in the L Intermediate of Bacteriorhodopsin: A Fourier-Transform Infrared Spectroscopic Study

Cited by 114 publications

References 48 publications

Water Structural Changes in the L and M Photocycle Intermediates of Bacteriorhodopsin as Revealed by Time-Resolved Step-Scan Fourier Transform Infrared (FTIR) Spectroscopy

Water Structural Changes in the L and M Photocycle Intermediates of Bacteriorhodopsin as Revealed by Time-Resolved Step-Scan Fourier Transform Infrared (FTIR) Spectroscopy

Factors That Differentiate the H-bond Strengths of Water Near the Schiff Bases in Bacteriorhodopsin and Anabaena Sensory Rhodopsin

The Photocycle of a Flavin-binding Domain of the Blue Light Photoreceptor Phototropin

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