Conformational Changes in the Core Structure of Bacteriorhodopsin

Kluge, Tilmann; Olejnik, Judith; Smilowitz, Laura; Rothschild, Kenneth J.

doi:10.1021/bi9802465

Cited by 24 publications

(31 citation statements)

References 58 publications

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“…This band is missing in the resonance Raman spectra of pR (40). In bacteriorhodopsin a similar band was attributed to an amide II response to isomerization (41). The appearance of a 1550 cm À1 band on a timescale on the order of tens of picoseconds was also observed for SRII (42) and halorhodopsin (43) and was interpreted as a relax- ation of protein vibrational bands, possibly amide II bands of the protein backbone, perturbed by the ultrafast retinal photoisomerization.…”

Section: Discussionmentioning

confidence: 82%

Characterization of the Primary Photochemistry of Proteorhodopsin with Femtosecond Spectroscopy

Rupenyan

Stokkum

Arents

et al. 2008

Biophysical Journal

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Proteorhodopsin is an ion-translocating member of the microbial rhodopsin family. Light absorption by its retinal chromophore initiates a photocycle, driven by trans/cis isomerization, leading to transmembrane translocation of a proton toward the extracellular side of the cytoplasmic membrane. Here we report a study on the photoisomerization dynamics of the retinal chromophore of proteorhodopsin, using femtosecond time-resolved spectroscopy, by probing in the visible- and in the midinfrared spectral regions. Experiments were performed both at pH 9.5 (a physiologically relevant pH value in which the primary proton acceptor of the protonated Schiff base, Asp(97), is deprotonated) and at pH 6.5 (with Asp(97) protonated). Simultaneous analysis of the data sets recorded in the two spectral regions and at both pH values reveals a multiexponential excited state decay, with time constants of approximately 0.2 ps, approximately 2 ps, and approximately 20 ps. From the difference spectra associated with these dynamics, we conclude that there are two chromophore-isomerization pathways that lead to the K-state: one with an effective rate of approximately (2 ps)(-1) and the other with a rate of approximately (20 ps)(-1). At high pH, both pathways are equally effective, with an estimated quantum yield for K-formation of approximately 0.7. At pH 6.5, the slower pathway is less productive, which results in an isomerization quantum yield of 0.5. We further observe an ultrafast response of residue Asp(227), which forms part of the counterion complex, corresponding to a strengthening of its hydrogen bond with the Schiff base on K-state formation; and a feature that develops on the 0.2 ps and 2 ps timescale and probably reflects a response of an amide II band in reaction to the isomerization process.

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Section: Discussionmentioning

confidence: 82%

Characterization of the Primary Photochemistry of Proteorhodopsin with Femtosecond Spectroscopy

Rupenyan

Stokkum

Arents

et al. 2008

Biophysical Journal

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“…A similar amide II downshift was observed in GPR (32) but only at room temperature on an ultrafast time scale. In contrast, in the low-temperature FTIR difference spectrum of BR, a pair of bands is assigned to the upshift of an amide II mode (41). …”

Section: Resultsmentioning

confidence: 99%

Different Structural Changes Occur in Blue- and Green-Proteorhodopsins during the Primary Photoreaction

et al. 2008

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We examine the structural changes during the primary photoreaction in blue-absorbing proteorhodopsin (BPR), a light-driven retinylidene proton pump, using low-temperature FTIR difference spectroscopy. Comparison of the light induced BPR difference spectrum recorded at 80 K to that of green-absorbing proteorhodopsin (GPR) reveals that there are several differences in the BPR and GPR primary photoreactions despite the similar structure of the retinal chromophore and all-trans → 13-cis isomerization. Strong bands near 1700 cm−1 assigned previously to a change in hydrogen bonding of Asn230 in GPR are still present in BPR but in addition bands in the same region are assigned on the basis of site-directed mutagenesis to changes occurring in Gln105. In the amide II region bands are assigned on the basis of total-N15 labeling to structural changes of the protein backbone, although no such bands were previously observed for GPR. A band at 3642 cm−1 in BPR, assigned to the OH stretching mode of a water molecule on the basis of H218O substitution, appears at a different frequency than a band at 3626 cm−1 previously assigned to a water molecule in GPR. However, the substitution of Gln105 for Leu105 in BPR leads to the appearance of both bands at 3642 and 3626 cm−1 indicating the waters assigned in BPR and GPR exist in separate distinct locations and can coexist in the GPR-like Q105L mutant of BPR. These results indicate that there exist significant differences in the conformational changes occurring in these two types proteorhodopsin during the initial photoreaction despite their similar chromophores structures, which might reflect a different arrangement of water in the active site as well as substitution of a hydrophilic for hydrophobic residue at residue 105.

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“…Earlier studies on bacteriorhodopsin indicated that internal, core domains of bacterio‐opsin are much more accessible for H 2 O/D 2 O exchange after bleaching than in the native pigment with the chromophore in place, indicating a more open and hydrophilic structure (24). However, no changes in the protonation states of buried aspartates Asp96 and Asp115 were detected upon reconstitution, just an IR frequency shift indicating a more hydrophobic environment in the reconstituted state (25).…”

Section: Discussionmentioning

confidence: 99%

Chromophore Interaction in Xanthorhodopsin—Retinal Dependence of Salinixanthin Binding^†

Imasheva

Balashov

Wang

et al. 2008

Photochem & Photobiology

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Xanthorhodopsin is a light-driven proton pump in the extremely halophilic bacterium Salinibacter ruber. Its unique feature is that besides retinal it has a carotenoid, salinixanthin, with a light harvesting function. Tight and specific binding of the carotenoid antenna is controlled by binding of the retinal. Addition of all-trans retinal to xanthorhodopsin bleached with hydroxylamine restores not only the retinal chromophore absorption band, but causes sharpening of the salinixanthin bands reflecting its rigid binding by the protein. In this report we examine the correlation of the changes in the two chromophores during bleaching and reconstitution with native all-trans retinal, artificial retinal analogues, and retinol. Bleaching and reconstitution both appear to be multi-stage processes. The carotenoid absorption changes during bleaching occurred not only uponhydrolysis of the Schiff base but continued while the retinal was leaving its binding site. In the case of reconstitution, the 13-desmethyl analogue formed the protonated Schiff base slower than retinal, and provided the opportunity to observe changes in carotenoid binding at various stages. The characteristic sharpening of the carotenoid bands, indicative of its reduced conformational heterogeneity in the binding site, occurs already when the retinal occupies the binding site but the covalent bond to Lys-227 via a Schiff base is not yet formed. This is confirmed by the results for retinol reconstitution, where the Schiff base does not form but the carotenoid exhibits its characteristic spectral change from the binding.

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Conformational Changes in the Core Structure of Bacteriorhodopsin

Cited by 24 publications

References 58 publications

Characterization of the Primary Photochemistry of Proteorhodopsin with Femtosecond Spectroscopy

Characterization of the Primary Photochemistry of Proteorhodopsin with Femtosecond Spectroscopy

Different Structural Changes Occur in Blue- and Green-Proteorhodopsins during the Primary Photoreaction

Chromophore Interaction in Xanthorhodopsin—Retinal Dependence of Salinixanthin Binding^†

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Conformational Changes in the Core Structure of Bacteriorhodopsin

Cited by 24 publications

References 58 publications

Characterization of the Primary Photochemistry of Proteorhodopsin with Femtosecond Spectroscopy

Characterization of the Primary Photochemistry of Proteorhodopsin with Femtosecond Spectroscopy

Different Structural Changes Occur in Blue- and Green-Proteorhodopsins during the Primary Photoreaction

Chromophore Interaction in Xanthorhodopsin—Retinal Dependence of Salinixanthin Binding†

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Chromophore Interaction in Xanthorhodopsin—Retinal Dependence of Salinixanthin Binding^†