Identification of retinal isomers isolated from bacteriorhodopsin

Pettei, Michael J.; Yudd, Anthony P.; Nakanishi, Koji; Henselman, Robert A.; Stoeckenius, Walther

doi:10.1021/bi00628a031

Cited by 241 publications

(173 citation statements)

References 15 publications

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“…5B and Table 1. The retinal composition of D96N bR is similar to that of WT bR, that is, almost equal amounts of the all-trans and 13-cis isoforms exist in the dark, whereas the all-trans form predominates (~93%) over 13-cis after light adaptation (Pettei et al, 1977). In contrast, D96N/W12I bR shows a reduction in the degree of difference in isomeric composition between the light-and dark-adapted states (64% all-trans and 32% 13-cis in the dark-adapted state; 68% all-trans, 23% 13-cis, and trace amounts of 9-cis and 11-cis in the light-adapted state).…”

Section: Hplc Analysis Of Retinal In D96n and D96n/w12i Mutantsmentioning

confidence: 85%

Role of trimer–trimer interaction of bacteriorhodopsin studied by optical spectroscopy and high-speed atomic force microscopy

Yamashita

Inoue

Shibata

et al. 2013

Journal of Structural Biology

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Bacteriorhodopsin (bR) trimers form a two-dimensional hexagonal lattice in the purple membrane of Halobacterium salinarum. However, the physiological significance of forming the lattice has long been elusive. Here, we study this issue by comparing properties of assembled and non-assembled bR trimers using directed mutagenesis, high-speed atomic force microscopy (HS-AFM), optical spectroscopy, and a proton pumping assay. First, we show that the bonds formed between W12 and F135 amino acid residues are responsible for trimer-trimer association that leads to lattice assembly; the lattice is completely disrupted in both W12I and F135I mutants. HS-AFM imaging reveals that both crystallized D96N and non-crystallized D96N/W12I mutants undergo a large conformational change (i.e., outward E-F loop displacement) upon light-activation. However, lattice disruption significantly reduces the rate of conformational change under continuous light illumination. Nevertheless, the quantum yield of M-state formation, measured by low-temperature UV-visible spectroscopy, and proton pumping efficiency are unaffected by lattice disruption. From these results, we conclude that trimer-trimer association plays essential roles in providing bound retinal with an appropriate environment to maintain its full photo-reactivity and in maintaining the natural photo-reaction pathway.

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Section: Hplc Analysis Of Retinal In D96n and D96n/w12i Mutantsmentioning

confidence: 85%

Role of trimer–trimer interaction of bacteriorhodopsin studied by optical spectroscopy and high-speed atomic force microscopy

Yamashita

Inoue

Shibata

et al. 2013

Journal of Structural Biology

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“…3 for a recent review). The light-adapted form of bR contains only all-trans retinal, whereas the dark-adapted form contains a 1:1 mixture of 13-cis and all-trans (4). Because of its long-term stability against thermal, chemical, and photochemical degradation and its desirable photoelectric and photochromic properties, bR has attracted much interest as a material for biooptics and bioelectronics (5).…”

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confidence: 99%

Bacteriorhodopsin (bR) as an electronic conduction medium: Current transport through bR-containing monolayers

Jin

Friedman²,

Sheves³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

157

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Studying electron transport (ET) through proteins is hampered by achieving reproducible experimental configurations, particularly electronic contacts to the proteins. The transmembrane protein bacteriorhodopsin (bR), a natural light-activated proton pump in purple membranes of Halobacterium salinarum, is well studied for biomolecular electronics because of its sturdiness over a wide range of conditions. To date, related studies of dry bR systems focused on photovoltage generation and photoconduction with multilayers, rather than on the ET ability of bR, which is understandable because ET across 5-nm-thick, apparently insulating membranes is not obvious. Here we show that electronic current passes through bR-containing artificial lipid bilayers in solid ''electrode-bilayer-electrode'' structures and that the current through the protein is more than four orders of magnitude higher than would be estimated for direct tunneling through 5-nm, water-free peptides. We find that ET occurs only if retinal or a close analogue is present in the protein. As long as the retinal can isomerize after light absorption, there is a photo-ET effect. The contribution of light-driven proton pumping to the steady-state photocurrents is negligible. Possible implications in view of the suggested early evolutionary origin of halobacteria are noted. molecular electronics ͉ vesicles ͉ bioelectronics B acteriorhodopsin (bR) is a protein-chromophore complex that serves as a light-driven proton pump in the purple membrane (PM) of Halobacterium salinarum (1). It has been shown that the protein is composed of seven transmembrane helices with a retinal chromophore covalently bound in the central region via a protonated Schiff base to a lysine residue (Fig. 1A). The PM is organized in a 2D hexagonal crystal lattice with a unit cell dimension of Ϸ6.2 nm. Electron crystallography has indicated that bR is organized into trimers in which lipids mediate intertrimer contact (2). Light absorption by bR initiates a multistep reaction cycle with several distinct spectroscopic intermediates: J 625 , K 590 , L 550 , M 412 , N 560 , and O 640 . More details on the molecular alterations that occur during the photocycle were recently obtained from x-ray diffraction studies (see ref. 3 for a recent review). The light-adapted form of bR contains only all-trans retinal, whereas the dark-adapted form contains a 1:1 mixture of 13-cis and all-trans (4). Because of its long-term stability against thermal, chemical, and photochemical degradation and its desirable photoelectric and photochromic properties, bR has attracted much interest as a material for biooptics and bioelectronics (5). Most of these efforts focused on multilayers and their photovoltage͞photocurrent generation (6-8) and photoconduction (9).In principle, PM patches (Ϸ5 nm thick, a few m in size; see Fig. 1B) can serve as a model protein material that is important for both planar junction fabrication and current transport measurements, because the 5-nm membrane is well beyond the thickness over which tu...

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“…The chromophoric part of native bR is derived from all-trans retinal and 13-cis retical in dark-adapted bR, and from all-trans retinal in light-adapted bR [2]. The chromophore is protonated Schiff-base bound to the lysine-216 residue [3,4].…”

mentioning

confidence: 99%

“…These modifications allow us to estimate the effect of the lack of the chromophoric six-membered ring and the influence of the steric and electronic factors due to this ring, the chromophoric 5,6 and 7,s double bond on bR. Continuing our studies on bR analogues with modified chromophores [ 12,131 we now report on the synthesis of retinal analogues [1][2][3][4][5]. the binding of their all-trans isomers to bacterioopsin and the light-driven proton pump capacity of the ensuing bR analogues.…”

mentioning

confidence: 99%

Bacteriorhodopsins with chromophores modified at the β‐ionone site

Muradin-Szweykowska¹,

Pardoen²,

Dobbelstein³

et al. 1984

European Journal of Biochemistry

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The binding to bacterioopsin of the all-trans isomers of retinal analogues lacking the six-membered ring and differing in length of the conjugated chain, as well as the light-driven action of the proton pump of the resulting bacteriorhodopsin analogues, were studied. The 'opsin shifts' in these modified bacteriorhodopsins are all around 2700 cm-' and do not depend on the number of double bonds in the chromophore. These experimental results suggest that the 4800 cm-' 'opsin shift' in unmodified bacteriorhodopsin consists of a contribution of about 2700 cm-' due to the interaction of the protonated Schiff-base with the counterion. The extra 2100cm-shift in bacteriorhodopsin is due to the specific interaction of the cyclohexene ring and the protein. Only the bacteriorhodopsin analogue with the same number of conjugated double bonds in the chromophore as bacteriorhodopsin itself shows light-driven proton pump action.Bacteriorhodopsin (hereafter bR), the only protein in the purple membrane of the halophilic microorganism Halobacterium halobium, occurs in a dark-adapted ( A, , , = 558 nm) and a light-adapted form ( A, , , = 568 nm), which are interconvertible [l]. The chromophoric part of native bR is derived from all-trans retinal and 13-cis retical in dark-adapted bR, and from all-trans retinal in light-adapted bR [2]. The chromophore is protonated Schiff-base bound to the lysine-216 residue [3,4]. Light-adapted bR functions as a light-driven proton pump, pumping protons from the inside to the outside of the bacterial cell thereby creating a proton gradient across the bacterial membrane to generate the energy for ATP synthesis [5 -81. The release of the chromophore from the bR binding site can be achieved without denaturation by irradiation of bR with visible light in the presence of hydroxylamine, leaving a chromophore-free apoprotein (called bacterioopsin) and all-trans retinal oxime [9]. Addition of all-trans retinal to bacterioopsin results in immediate formation of the so called 430 -460-nm intermediate complex which rapidly converts into all-trans bR [lo] (A, , , = 568 nm). This regenerated bR, after incorporation into phospholipid vesicles pumps protons upon illumination as efficiently as native bR [6]. The red shift in the absorption maximum of dark-adapted bR (in cm-') relative to the protonated all-trans retinal Schiff-base model compound (in cm-', A,,,=440 nm), which is called 'opsin shift' [ll], is believed to be caused by special proteinchromophore interactions. Contributing factors for this 'opsin shift' are an interaction of an anion located near the cyclohexene part of the chromophore in addition to the interaction of the counterion with the protonated Schiff-base part [I I].In order to study the importance of the six-membered ring and of the 5,6 and 7,8 double bond in the chromophore for the purple colour and proton pump activity of bR, our straAbbreviations. bR, bacteriorhodopsin; bR(1) -bR(5), bacteriorhodopsin containing the retinal analogues 1 -5. tegy was to investigate the binding to bacterioo...

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Identification of retinal isomers isolated from bacteriorhodopsin

Cited by 241 publications

References 15 publications

Role of trimer–trimer interaction of bacteriorhodopsin studied by optical spectroscopy and high-speed atomic force microscopy

Role of trimer–trimer interaction of bacteriorhodopsin studied by optical spectroscopy and high-speed atomic force microscopy

Bacteriorhodopsin (bR) as an electronic conduction medium: Current transport through bR-containing monolayers

Bacteriorhodopsins with chromophores modified at the β‐ionone site

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