ChemInform Abstract: Mechanism for the Opsin Shift of Retinal′s Absorption in Bacteriorhodopsin.

Lugtenburg, J.; Muradin-Szweykowska, M.; Heeremans, C. E. M.; Pardoen, J. A.; Harbison, Gerard S.; Herzfeld, Judith; Griffin, Robert G.; Smith, Steven O.; Mathies, Richard A.

doi:10.1002/chin.198640067

Cited by 26 publications

(29 citation statements)

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“…Previous FTIR (Rothschild & Marrero, 1982;Bagley et al, 1982), laser Raman (Smith et al, 1987;Hildebrandt & Stockburger, 1984), and NMR studies (Harbison et al, 1983(Harbison et al, , 1985a, together with chemical analogue experiments (Nakanishi et al, 1980;Lugtenburg et al, 1986;Spudich et al, 1986) and theoretical calculations (Honig et al, 1976), have contributed to our present understanding of the electronic properties of the chromophore in bR. It appears that several mechanisms contribute to the opsin shift, the three most important being the configuration of the 6-7 (6-s) single bond, which influences the extension of the 7r-electron system into the the p-ionone ring, the strength of the hydrogen bond at the Schiff base, and electrostatic interactions with other nearby protein charges [see, e.g., Lugtenburg et al (1986) and references cited therein].…”

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“…It appears that several mechanisms contribute to the opsin shift, the three most important being the configuration of the 6-7 (6-s) single bond, which influences the extension of the 7r-electron system into the the p-ionone ring, the strength of the hydrogen bond at the Schiff base, and electrostatic interactions with other nearby protein charges [see, e.g., Lugtenburg et al (1986) and references cited therein]. The importance of the hydrogen bonding of the Schiff base may also be appreciated by comparing the pK, > 12 of the Schiff base in bR (Ehrenberg et al, 1980;Doukas et al, 1981;Druckmann et al, 1982) with the pKa N 7 typically observed for protonated Schiff base model compounds (Favrot et al, 1978).…”

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Nuclear magnetic resonance study of the Schiff base in bacteriorhodopsin: counterion effects on the nitrogen-15 shift anisotropy

Groot¹,

Harbison²,

Herzfeld³

et al. 1989

Biochemistry

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High-resolution, solid-state 15N NMR has been used to study the chemical shift anisotropies of the Schiff bases in bacteriorhodopsin (bR) and in an extensive series of model compounds. Using slow-spinning techniques, we are able to obtain sufficient rotational sideband intensity to determine the full 15N chemical shift anisotropy for the Schiff base nitrogen in bR548 and bR568. Comparisons are made between all-trans-bR568 and N-all-trans-retinylidene butylimine salts with halide, phenolate, and carboxylate counterions. It is argued that for the model compounds the variation in 15N chemical shift reflects the variation in (hydrogen) bond strength with the various counterions. The results suggest that carboxylates and tyrosinates may form hydrogen bonds of comparable strength in a hydrophobic environment. Thus, the hydrogen bonding strength of a counterion depends on factors that are not completely reflected in the solution pKa of its conjugate acid. For the model compounds, the two most downfield principal values of the 15N chemical shift tensor, sigma 22 and sigma 33, vary dramatically with different counterions, whereas sigma 11 remains essentially unaffected. In addition, there exists a linear correlation between sigma 22 and sigma 33, which suggests that a single mechanism is responsible for the variation in chemical shifts present in all three classes of model compounds. The data for bR568 follow this trend, but the isotropic shift is 11 ppm further upfield than any of the model compounds. This extreme value suggests an unusually weak hydrogen bond in the protein.

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Nuclear magnetic resonance study of the Schiff base in bacteriorhodopsin: counterion effects on the nitrogen-15 shift anisotropy

Groot¹,

Harbison²,

Herzfeld³

et al. 1989

Biochemistry

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“…dark-adapted bR comprising a mixture of bR,,, and bR568 (Scherrer et al, 1989); bR,,,, 1 3 4 s component of dark-adapted bR; bR568, light-adapted bR; bR,,, blue membrane obtained by acidification or deionization of bR,,; bR!65, acid purple membrane obtained by acidification of blue membrane with HCI; CP, cross-polarization;A fraction of total intensity in the difference spectrum; FWHM, full width at half-maximum; MAS, magic angle spinning; PSB, protonated Schiff base; PM, purple membrane; ppm, parts per million; TMS, tetramethylsilane; A, , , , wavelength of maximum visible absorption; wr, magic angle spinning speed; u, chemical shift. There have been numerous investigations of the opsin shift (Honig et al, 1976;Nakanishi et al, 1980;Bagley et al, 1982;Rothschild & Marrero, 1982;Harbison et al, 1983Harbison et al, , 1985aHildebrandt & Stockburger, 1984;Lugtenburg et al, 1986;Spudich et al, 1986;Smith et al, 1987), and currently, the effect is thought to be due to a combination of various chromophore-protein interactions (Muradin-Szweykowska et al, 1984;Lugtenburg et al, 1986). Important contributions include a weak hydrogen bond between the protonated Schiff base and its counterion (Oseroff & Callender, 1974;Blatz & Mohler, 1975;Harbison et al, 1983;de Groot et al, 1989) and isomerization of the C6-C7 bond from the 6-s-cis structure dominating in solution to the 6-s-trans bR geometry illustrated in Figure 1 (Harbison et al,1 98 5a,b).…”

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Solid-state carbon-13 and nitrogen-15 NMR study of the low pH forms of bacteriorhodopsin

Groot¹,

Smith²,

Courtin³

et al. 1990

Biochemistry

135

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The visible absorption of bacteriorhodopsin (bR) is highly sensitive to pH, the maximum shifting from 568 nm (pH 7) to approximately 600 nm (pH 2) and back to 565 nm (pH 0) as the pH is decreased further with HCl. Blue membrane (lambda max greater than 600 nm) is also formed by deionization of neutral purple membrane suspensions. Low-temperature, magic angle spinning 13C and 15N NMR was used to investigate the transitions to the blue and acid purple states. The 15N NMR studies involved [epsilon-15N]lysine bR, allowing a detailed investigation of effects at the Schiff base nitrogen. The 15N resonance shifts approximately 16 ppm upfield in the neutral purple to blue transition and returns to its original value in the blue to acid purple transition. Thus, the 15N shift correlates directly with the color changes, suggesting an important contribution of the Schiff base counterion to the "opsin shift". The results indicate weaker hydrogen bonding in the blue form than in the two purple forms and permit a determination of the contribution of the weak hydrogen bonding to the opsin shift at a neutral pH of approximately 2000 cm-1. An explanation of the mechanism of the purple to blue to purple transition is given in terms of the complex counterion model. The 13C NMR experiments were performed on samples specifically 13C labeled at the C-5, C-12, C-13, C-14, or C-15 positions in the retinylidene chromophore. The effects of the purple to blue to purple transitions on the isotropic chemical shifts for the various 13C resonances are relatively small. It appears that bR600 consists of at least four different species. The data confirm the presence of 13-cis- and all-trans-retinal in the blue form, as in neutral purple dark-adapted bR. All spectra of the blue and acid purple bR show substantial inhomogeneous broadening which indicates additional irregular distortions of the protein lattice. The amount of distortion correlates with the variation of the pH, and not with the color change.

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“…In this case, the substitution at the retinal positions 4 and 18 (methyl group at position 5 ) or the introduction of the a-configuration of the ring double bond led to strong A,,, shifts of the absorption maxima [8, 32 -341. These absorption changes, however, can be explained by a direct influence of the substitutions on the interactions between the chromophore and a proposed ion pair located in the ring region (for a proposed arrangement of the ion pair formed by two amino acid residues see [12,321). For the shifts of the absorption maximum of 13-methoxy BR, a disturbed interaction with the protein around the region of the Schiff base group must be assumed, as supported by the results obtained with 13-ethyl retinal BR.…”

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Methoxyretinals in bacteriorhodopsin

Gärtner¹,

Oesterhelt

1988

European Journal of Biochemistry

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Analogue bacteriorhodopsins (BRs) were reconstituted from bacterioopsin and 9-, 11 -, or 13-methoxyretinals or their demethyl derivatives, respectively. In organic solvents the retinals occur as cis isomers of the respective double bonds carrying the methoxy group. 9-Methoxyretinal, present as the 9-cis isomer, does not form an analogue BR with bacterioopsin in the dark. Upon illumination, a BR is produced with an absorbance maximum at 560 nm. This compound is thermally unstable, and converts back into the 9-cis-containing complex (Amax = 410 nm) in the dark. Removal of the 13-methyl group from this compound (= 9-methoxy 13-demethyl retinal) does not change the 9-cis configuration of the free retinal, but allows the reconstitution of a thermally stable chromoprotein absorbing around 500 nm with a proton translocation rate of about 10% of the BR value, comparable to the 13-demethyl BR value [Gartner, W., Towner, P., Hopf, H. & Oesterhelt, D. (1983) Biochemistry 22, 2637 -26441. 11-Methoxy BRs (13-demethyl and 9,13-didemethyl) absorb around 530 nm and are inactive. 13-Methoxy retinal (13-cis isomer) reconstitutes a chromoprotein with an absorbance maximum at 51 5 nm, which can be photoconverted to a thermostable 460-nm-absorbing complex, For the 51 5-nm-absorbing species of 13-methoxy BR a light-induced proton translocation was not detected in measurements with cell vesicles (detection of pH changes in the vesicle preparation). Only by photocurrent measurements in a bilayer experiment could a very diminished photocurrent be detected, about 3 -2% of BR, [Fendler et al. (1987) Biochim. Biophys. Acta 893, 60-681. The reconstitution rate of 13-methoxy BR from 13-methoxy retinal and bacterioopsin is slower by a factor of 40 compared to 13-ethyl BR, although both substituents are of similar size.The position 13 of retinal was found to be most sensitive for regulation of the absorption maximum and the formation and stability of the all-trans isomer, which is the active form for light-induced proton translocation. The results suggest that an electronic interaction with a charged residue of the binding site exists around position 13 of retinal, which is disturbed when a methoxy group replaces the methyl or ethyl group at that position. This electronic interaction is essential for maintaining the active all-trans configuration of retinal.Halohacterium halohium is able to grow phototrophically under oxygen-deprived conditions [l]. This phenomenon is based on the membrane-embedded protein bacteriorhodopsin (BR), which upon illumination functions as a light-driven proton pump (for a comprehensive review see [2]). Essential for this process is the prosthetic group of BR, a molecule of retinal (i), which is covalently bound via a protonated Schiff base (PSBH', 2) to a lysine residue of the polypeptide chain (see Fig. 1). The resting state of BR, referred to as the dark adapted form (BRDA), contains a nearly equal distribution o f the all-trans and the 1 3 4 s isomer of retinal [3]. Upon illumination, both isomeric forms of retin...

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ChemInform Abstract: Mechanism for the Opsin Shift of Retinal′s Absorption in Bacteriorhodopsin.

Cited by 26 publications

References 1 publication

Nuclear magnetic resonance study of the Schiff base in bacteriorhodopsin: counterion effects on the nitrogen-15 shift anisotropy

Nuclear magnetic resonance study of the Schiff base in bacteriorhodopsin: counterion effects on the nitrogen-15 shift anisotropy

Solid-state carbon-13 and nitrogen-15 NMR study of the low pH forms of bacteriorhodopsin

Methoxyretinals in bacteriorhodopsin

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