Synthetic retinals as probes for the binding site and photoreactions in rhodopsins

Ottolenghi, Michael; Sheves, Mordechai

doi:10.1007/bf01870951

Cited by 78 publications

(62 citation statements)

References 142 publications

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“…The synthesis of some of the retinal analogues has been described previously 46 . Details of the synthesis of compounds II-IX, as well as 1 H NMR, 13 C NMR and HMRS data, can be found in Supplementary Methods.…”

Section: Methodsmentioning

confidence: 99%

“…Reconstitution of Arch3 and four ChR variants with A2 retinal, which carries an extra double bond, resulted in functional pigments with enhanced longwavelength sensitivity 29 . Extensive work on bacteriorhodopsin (BR) [45][46][47][48][49] using synthetic retinal analogues suggests that many other retinal analogues, particularly those with ring modifications, might be applicable for the purpose of chromophore substitution. However, in most cases, it was only shown that the absorption maximum was shifted, without analysing the rhodopsin function.…”

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confidence: 99%

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Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

et al. 2014

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Optogenetic tools have become indispensable in neuroscience to stimulate or inhibit excitable cells by light. Channelrhodopsin-2 (ChR2) variants have been established by mutating the opsin backbone or by mining related algal genomes. As an alternative strategy, we surveyed synthetic retinal analogues combined with microbial rhodopsins for functional and spectral properties, capitalizing on assays in C. elegans, HEK cells and larval Drosophila. Compared with all-trans retinal (ATR), Dimethylamino-retinal (DMAR) shifts the action spectra maxima of ChR2 variants H134R and H134R/T159C from 480 to 520 nm. Moreover, DMAR decelerates the photocycle of ChR2(H134R) and (H134R/T159C), thereby reducing the light intensity required for persistent channel activation. In hyperpolarizing archaerhodopsin-3 and Mac, naphthyl-retinal and thiophene-retinal support activity alike ATR, yet at altered peak wavelengths. Our experiments enable applications of retinal analogues in colour tuning and altering photocycle characteristics of optogenetic tools, thereby increasing the operational light sensitivity of existing cell lines or transgenic animals.

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

et al. 2014

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“…Two Titratable Groups Control BPR Absorption SpectraOne of the most notable distinguishing properties of retinal as a chromophore is the large variation of its absorption spectrum depending on interaction with the apoprotein ("spectral tuning") (13,14). In rhodopsins, retinal is covalently attached to the ⑀-amino group of a lysine residue, forming a protonated retinylidene Schiff base.…”

Section: Resultsmentioning

confidence: 99%

Spectroscopic and Photochemical Characterization of a Deep Ocean Proteorhodopsin

Wang

Sineshchekov

Spudich

et al. 2003

Journal of Biological Chemistry

159

270

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A second group of proteorhodopsin-encoding genes (blue-absorbing proteorhodopsin, BPR) differing by 20 -30% in predicted primary structure from the first-discovered green-absorbing (GPR) group has been detected in picoplankton from Hawaiian deep sea water. Here we compare BPR and GPR absorption spectra, photochemical reactions, and proton transport activity. The photochemical reaction cycle of Hawaiian deep ocean BPR in cells is 10-fold slower than that of GPR with very low accumulation of a deprotonated Schiff base intermediate in cells and exhibits mechanistic differences, some of which are due to its glutamine residue rather than leucine at position 105. In contrast to GPR and other characterized microbial rhodopsins, spectral titrations of BPR indicate that a second titratable group, in addition to the retinylidene Schiff base counterion Asp-97, modulates the absorption spectrum near neutral pH. Mutant analysis confirms that Asp-97 and Glu-108 are proton acceptor and proton donor, respectively, in retinylidene Schiff base proton transfer reactions during the BPR photocycle as previously shown for GPR, but BPR contains an alternative acceptor evident in its D97N mutant, possibly the same as the second titratable group modulating the absorption spectrum. BPR, similar to GPR, carries out outward light-driven proton transport in Escherichia coli vesicles but with a reduced translocation rate attributable to its slower photocycle. In energized E. coli cells at physiological pH, the net effect of BPR photocycling is to generate proton currents dominated by a triggered proton influx, rather than efflux as observed with GPR-containing cells. Reversal of the proton current with the K ؉ -ionophore valinomycin supports that the influx is because of voltagegated channels in the E. coli cell membrane. These observations demonstrate diversity in photochemistry and mechanism among proteorhodopsins. Calculations of photon fluence rates at different ocean depths show that the difference in photocycle rates between GPR and BPR as well as their different absorption maxima may be explained as an adaptation to the different light intensities available in their respective marine environments. Finally, the results raise the possibility of regulatory (i.e. sensory) rather than energy harvesting functions of some members of the proteorhodopsin family.

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“…We approached this issue by combining the lightinduced AFS technique with the well-established methodology of replacing the native retinylidene chromophore of bR with synthetic analogs (for a review, see ref. 29). The specific choices were artificial pigments (I, II, and III) formed from three locked retinal analogs 1,2 (30) and 3 (31,32), in which the critical C 13 AC 14 isomerization is blocked by a 5-membered ring structure.…”

Section: Resultsmentioning

confidence: 99%

Microsecond atomic force sensing of protein conformational dynamics: Implications for the primary light-induced events in bacteriorhodopsin

Rousso

Khachatryan²,

Gat³

et al. 1997

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

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In this paper a new atomic force sensing technique is presented for dynamically probing conformational changes in proteins. The method is applied to the light-induced changes in the membrane-bound proton pump bacteriorhodopsin (bR). The microsecond time-resolution of the method, as presently implemented, covers many of the intermediates of the bR photocycle which is well characterized by spectroscopical methods. In addition to the native pigment, we have studied bR proteins substituted with chemically modified retinal chromophores. These synthetic chromophores were designed to restrict their ability to isomerize, while maintaining the basic characteristic of a large lightinduced charge redistribution in the vertically excited Franck--Condon state. An analysis of the atomic force sensing signals lead us to conclude that protein conformational changes in bR can be initiated as a result of a light-triggered redistribution of electronic charge in the retinal chromophore, even when isomerization cannot take place. Although the coupling mechanism of such changes to the light-induced proton pump is still not established, our data question the current working hypothesis which attributes all primary events in retinal proteins to an initial transNcis isomerization.

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Synthetic retinals as probes for the binding site and photoreactions in rhodopsins

Cited by 78 publications

References 142 publications

Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

Synthetic retinal analogues modify the spectral and kinetic characteristics of microbial rhodopsin optogenetic tools

Spectroscopic and Photochemical Characterization of a Deep Ocean Proteorhodopsin

Microsecond atomic force sensing of protein conformational dynamics: Implications for the primary light-induced events in bacteriorhodopsin

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