Retinal Isomerization Research Articles

PHOTOCHEMISTRY OF ALL‐TRANS‐ AND 13‐CIS‐14,15‐DIDEUTERIORETINAL

Abstract— All‐trans‐ and 13‐cis‐14,15‐dideuterioretinal were synthesized and their solution photochemistry examined. Quantum yields of trans→cis or cis→ trans photoisomerization and the number and ratio of primary photoproducts, determined by high pressure liquid chromatographic analysis, are essentially identical to that of their corresponding retinal isomer. The C‐14, C‐15 carbon‐hydrogen modes play no particularly important role in the deactivation of electronically excited linear polyenes related to retinal.

Light-induced reaction of halorhodopsin prepared under low salt conditions.

1. Membrane fraction containing halorhodopsin was prepared from the lysate of a mutant strain of Halobacterium halobium, Y1, which is defective in bacteriorhodopsin synthesis. 2. Irradiation of the membrane with red light at 0 degrees C decreased the absorbance intensities over the whole range of 500--600 nm and a new absorption peak appeared at about 400 nm. This process was reversed either by irradiation with blue light or simply by incubation in the dark. The wavelength at which the red light-induced absorbance decrease became maximum was 566 nm in the absence of NaCl and moved to 576 nm upon addition of NaCl. The midpoint for the conversion was at about 0.1 M NaCl. 3. Even though the membrane containing halorhodopsin was prepared in the absence of NaCl, this pigment was photochemically active in the sense that it could form a bathochromic photoproduct, a batho-intermediate, at -196 degrees C. 4. Retinal isomer composition during irradiation with red light at 0 degrees C was determined. Unirradiated halorhodopsin had predominantly all-trans retinal. With the increase of the blue-shifted product on irradiation, 13-cis content increased.

Retinal-protein interactions in bacteriorhodopsin monomers, dispersed in the detergent L-1690

Bacteriorhodopsin monomer dispersed in a solution of the detergent L-1690 could maintain the specific interaction between retinal and protein in the pH range 9.0-0.0 at 25°C. λ max of the absorbance spectrum was 550 nm at pH 9.0, 556 nm at pH 5.5, 609 nm at pH 2.1 and 570 nm at pH 0.0. Increasing the NaCl concentration in the solution promoted formation of the 609 nm product at pH 5.0-3.0 and also its transition to the 570 nm product at pH 2.5-1.0. Retinal isomer analysis gave a ratio of 13- cis- to all- trans-retinal of 53 : 47 at pH 5.5. When the pH of the solution was reduced, the relative content of all- trans-retinal increased and the ratio of 13- cis- to all- trans-retinal was 14 : 86 at pH 0.0. Illumination of the solution at pH 7.2 yielded a product containing 9- cis-retinal or 9- cis, 13- cis-retinal, which may be due to a reaction other than the photoreaction cycle.

Mediation of retinal photoisomerization by adduct formation with tris(6,6,7,7,8,8,8-heptafluoro-2,2-dimethyl-3,5-octanedionato)-europium(II).

Several retinal isomers are shown to form adducts in isooctane solution with a lanthanide beta-diketonate complex, tris(6,6,7,8,8,8-heptafluoro-2,2-dimethyl-3,5-octanedionato)europium(III) [Eu(fod)3]. The adducts are characterized by an absorption band whose lambda max at approximately 420 nm is red-shifted by approximately 60 nm from the lowest-energy absorption band in the free retinal isomers. Irradiation into this adduct band leads to photoisomerization. For example, photolysis of a 1 mM all-trans-retinal/3 mM Eu(fod)3 isooctane solution at 514.5 nm leads, with reasonable quantum efficiency (phi greater than or equal to 0.05), to isomeric mixtures that are considerably different from these produced with UV excitation in the absence of Eu(fod)3. Particularly noteworthy for a photolysis conducted in a nonpolar solvent is the presence of an appreciable quantity of 11-cis-retinal in the adduct photolysate.

Specific photoisomerization of retinal in squid rhodopsin and metarhodopsin

The composition of retinal isomers in the photosteady-state mixtures formed from squid rhodopsin and metarhodopsin was determined by high-pressure liquid chromatography. A large amount of 9- cis-retinal was obtained at liquid N 2 temperature when rhodopsin was irradiated with orange light, but only small quantities of 9- cis-retinal were obtained at 15°C. Scarcely any 9- cis-retinal was produced from metarhodopsin by irradiation at liquid N 2 temperature. A large quantity of 7- cis-retinal was found in the photoproduct of rhodopsin irradiated at solid carbon dioxide temperature, but not at 15°C and liquid N 2 temperature. 7- cis-Retinal was not produced from metarhodopsin at any temperatures. These results indicate that the photoisomerization of retinal is regulated by the structure of the retinal-binding site of this protein. The formation of 9- cis- and 7- cis-retinals is forbidden in the metarhodopsin protein.

Resonance Raman evidence for an all-trans to 13-cis isomerization in the proton-pumping cycle of bacteriorhodopsin.

Using a dual-beam flow technique, we have obtained resonance Raman spectra of the M412 photointermediates of both native purple membrane (15H M412) and purple membrane regenerated with 15-deuterioretinal (15D M412). For comparison, we have also obtained Raman spectra of the n-butylamine Schiff bases of the 13-cis and all-trans isomers of 15H and 15D retinal. The 15D model compound spectra, when compared to the 15H spectra, show isotopically induced spectral changes that are markedly different for the two isomers. There is a very close agreement between the frequency and intensity changes which occur upon deuteration of M412 and those which occur upon deuteration of the 13-cis model compound, but not even a qualitative correspondence exists when M412 and the all-trans model compound are similarly compared. These data demonstrate that the chromophore of M412 is an unprotonated Schiff base of 13-cis-retinal rather than all-trans-retinal. An analogous spectral comparison of 15H and 15D light-adapted bacteriorhodopsin (bRLA) with the 15H and 15D protonated Schiff bases of 13-cis- and all-trans-retinal demonstrates that bRLA contains an all-trans chromophore, in agreement with previous extraction experiments. Thus, a trans leads to cis isomerization occurs in the proton-pumping photocycle of Halobacterium halobium.

Formation of 9-cis- and 11-cis-retinal pigments from bacteriorhodopsin by irradiating purple membrane in acid.

Both light-adapted and dark-adapted forms of bacteriorhodopsin in purple membrane in 67% glycerol solution were allowed to stand in acidic conditions by the addition of HCl to final concentrations from 4 X 10(-4) to 2 X 10(-2) M for 24 h at 3 degrees C. Over this concentration range, the acid-induced products from both species showed a maximum absorbance around 600 nm and high-performance liquid chromatography of extracted retinal isomers revealed that the acid-induced form of bacteriorhodopsin has 13-cis- and all-trans-retinals in a molar ratio of 4:6, which is intermediate between those of the dark-adapted and the light-adapted forms at neutral pH values. Exposure of the acid-induced form of bacteriorhodopsin to light at wavelengths longer than 670 nm at 3 degrees C caused a decrease of the absorbance around 600 nm with a concomitant rise of the absorbance around 500 nm. The extract from the irradiated products of bacteriorhodopsin in acid contained 9-cis- and 11-cis-retinals in addition to 13-cis- and all-trans-retinals. The absorbance maximum estimated from the analysis of the absorption spectra and the composition of the isomers was found at 495 nm for the 9-cis-retinal pigment and around 560 nm for the 11-cis-retinal pigment. On irradiation with 438-nm light, the 9-cis-retinal pigment disappeared with a concomitant increase of both the 13-cis- and all-trans-retinal pigments as judged by chromophore analysis and the absorption spectrum. The 9-cis-retinal pigment brought to pH 9 exhibited a maximum absorbance at 450 nm; this could be decomposed by the action of hydroxylamine or converted to a form resembling normal bacteriorhidopsin by 438-nm irradiation.

ChemInform Abstract: ELECTRONIC AND VIBRATIONAL SPECTRAL INVESTIGATION OF THE MOLECULAR ASSOCIATION OF THE ALL‐TRANS ISOMERS OF RETINAL, RETINOL, AND RETINOIC ACID

Chemischer InformationsdienstVolume 11, Issue 30 Physical Organic Chemistry ChemInform Abstract: ELECTRONIC AND VIBRATIONAL SPECTRAL INVESTIGATION OF THE MOLECULAR ASSOCIATION OF THE ALL-TRANS ISOMERS OF RETINAL, RETINOL, AND RETINOIC ACID K. CHIHARA, K. CHIHARASearch for more papers by this authorW. H. WADDELL, W. H. WADDELLSearch for more papers by this author K. CHIHARA, K. CHIHARASearch for more papers by this authorW. H. WADDELL, W. H. WADDELLSearch for more papers by this author First published: July 29, 1980 https://doi.org/10.1002/chin.198030048AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume11, Issue30July 29, 1980 RelatedInformation

Longitudinal restrictions of the binding site of opsin as measured with retinal isomers and analogs

Longitudinal restrictions of the binding site of opsin as measured with retinal isomers and analogs

High-performance liquid chromatographic analysis of cis-trans stereoisomeric 3-dehydroretinals in the presence of retinal isomers

Summary A high-performance liquid chromatographic method for the separation of geometric isomers of 3-dehydroretinal in the presence of retinal isomers has been devised. Simultaneous determination of 11- cis -retinal and 11- cis -3-dehydroretinal was also achieved. Cis-trans photoisomerization of 3-dehydroretinal and retinal under the same conditions was examined and the possible importance of the 9- ci s isomer in the former polyenal series is considered.

The effect of retinal isomers on the VER and ERG of vitamin A deprived rats

Male weanling Long-Evans and Sprague-Dawley rats were administered a vitamin A deficient diet and the electroretinogram (ERG) and/or visual evoked response (VER) were recorded weekly. After 6–10 weeks, when the animal showed a greatly decreased VER and ERG, the animal was injected intraperitoneally with a retinal isomer. Following dark adaptation for 18 hr, the ERG and VER were recorded. The rat was then sacrificed and the retina dissected. The retina was used for either solubilizing the visual pigment in 2% Ammonyx LO or extracting the retinals in methylene chloride. The visual pigments were characterized by the absorbance spectra and the retinal isomers were identified by high pressure liquid Chromatography (HPLC). The physiologically occurring isomers of retinal (11- cis and all- trans) are incorporated in the retina of deprived animals and the VER and ERG amplitudes for a given light intensity, diminished as a result of vitamin A deprivation, are restored to their normal values. 9- cis retinal is also incorporated into the retina, as shown by extraction of the retina with organic solvent and identification of the isomers by HPLC; a pigment is formed which has a λ max blue-shifted from that of rhodopsin. The ERG and VER amplitudes of deprived rats injected with 9- cis retinal are restored to normal. No incorporation of 13- cis retinal into the retina is observed and the ERG and VER amplitudes are not significantly affected.

28] High-pressure liquid chromatography of vitamin A metabolites and analogs

Publisher Summary This chapter discusses high-pressure liquid chromatography (HPLC) of vitamin A metabolites and analogs. The application of HPLC to the separation of retinoids has revolutionized vitamin A chromatography and has proved to be an extremely powerful and valuable method for studying vitamin A metabolism. The use of HPLC offers several advantages over conventional methods of vitamin A chromatography, including rapid separation, excellent resolution, quantitative recovery, elimination of artifacts, and the capability of chromatographing a wide range of vitamin A compounds with only minor manipulation of the chromatographic solvent. Straight-phase HPLC on microparticulate silica gel columns has been applied to the study of retinol storage and distribution, retinal isomers and analogs, determination of serum retinol and retinoic acid, and isolation of urinary and fecal metabolites of retinoic acid. This chapter describes the high-pressure liquid chromatography methods developed and used in the authors' laboratory for separation of vitamin A metabolites and analogs. Straight-phase and reverse-phase high-pressure liquid chromatography methods are presented and their application to biochemical problems in the vitamin A field are discussed.

The Temperature Dependence of Carbon-13 Chemical Shifts of Retinal Isomers and Their Related Compounds

Abstract The temperature dependence of carbon-13 chemical shifts has been studied for retinal isomers, N-all-trans-retinylidenebutylamine(all-trans-NRB) and N-all-trans-retinylidenebutylammonium chloride (all-trans-NRB·HCl). The change in the chemical shift was discussed in terms of the temperature dependences of the polarizability and conformation of the polyene-chain. In 11-cis-retinal, the coplanarity between the two parts of polyene-chain separated by the cis bond increases with a lowering of the temperature. The equilibrium between 12-s-cis and 12-s-trans conformers in 11-cis-retinal was studied and it was shown the latter conformer is preferable at low temperatures. In all-trans-NRB and all-trans-NRB·HCl, the temperature dependences of the chemical shifts are similar to those in all-trans-retinal. The extents of the chemical shift changes are in the following order: NRB·HCl<NRB<retinal. This trend can be correlated with the order of the red shifts of the absorption spectra.

Isolation and characterization of a water-soluble photopigment from honeybee compound eye

By preparative electrophoresis on polyacrylamide gel, a water-soluble light-sensitive pigment was isolated from honeybee heads incubated with tritiated vitamin A. Thin layer chromatography (TLC) on silica gel demonstrated the presence of retinal isomers as a chromophore of the bee pigment. It seems that all- trans-retinal is bound to the protein and an isomerization to 11- cis-retinal occurs in the light. Preliminary spectrophotometric measurements show that illumination of the extracted pigment is followed by a decrease in absorbance with a maximum at about 440 nm. A Schiff's base linkage is suggested for the binding between retinal and protein. Comparisons with visual pigments from other insects and with squid retinochrome are discussed, together with the possibility that the bee pigment could be involved in the turnover of the visual pigment of the honeybee eye.

Sensory Transduction in Halobacterium halobium: Retinal Protein Pigment Controls UV-Induced Behavioral Response

Abstract Both photosystems, PS 370 and PS 565, controlling behavioral responses in Halobacterium halobium [E. Hildebrand and N. Dencher, Nature 257, 46 - 48 (1975)] are reversibly inhibited when bacteria are grown in the presence of 1 mM nicotine which is known to block biosynthesis of reti­nal. Photobehavior can be restored within som e minutes to hours by adding retinal to nicotine-treated bacteria, PS 370 thereby reappearing earlier than PS 565. The reconstitution rate depends on the concentration and on the kind of retinal isomers applied. All-trans retinal is most effective. PS 370 becomes fully sensitive if reconstituted in the presence of nicotine. This rules out the possi­bility that the alkaloid may directly inhibit steps of signal transmission following photoreception. The action spectrum of PS 370 regenerated with retinal alone of H. h., strain R1L3 (a mutant deficient in carotenoids), fails to show all secondary peaks around 450 nm which in strain R1 occur besides the prominent maximum at 370 nm. Addition of carotenoids (mainly α-bacterioruberin) to reconstituted cells of R1L3 restores the sensitivity in that spectral region. Carotenoids or flavin solely added to nicotine-treated bacteria cannot restore photobehavior. We conclude that the active pigment o f PS 370, which mediates the photophobic response to increase of light intensity (step-up response), represents a retinal protein complex and that carote­noids participate in photoreceptor function as accessory pigments. The biochemical relation of the UV-absorbing retinal protein complex to bacteriorhodopsin is discussed.

Spectroscopic properties of molecules related to hindered isomers of retinal

Proton and12C NMR study of molecules related to retinal has been carried out. The characteristic differences in spectral behaviour among 7-trans and 7-cis isomers have been established which would be useful in determining the structure of new isomers and identifying components in a mixture. Through coupling constant measurements and DNMR study it is clearly established that 7-cis isomers of β-ionyl derivatives and in turn 7-cis isomers of retinyl derivatives prefer a non-planar arrangement and this non-planarity brings about resonance destabilisation.

Time-resolved resonance Raman characterization of the bL550 intermediate and the two dark-adapted bRDA/560 forms of bacteriorhodopsin

The resonance Raman spectrum of the second intermediate in the bacteriorhodopsin cycle, bL550, is obtained by a simple flow technique. The Schiff base linkage in this intermediate appears to be protonated, contrary to previous suggestion. The fingerprint region of the spectrum of bL550 does not closely match those of any presently available model Schiff bases of retinal isomers, though some comparisons can be made. The resonance Raman spectrum of dark-adapted bacteriorhodopsin is obtained and decomposed by computer subtraction of the spectrum of bR570. The remaining spectrum does not match the spectra of any model compounds presently in the literature. The spectra of bL550 and dark-adapted bRDA/560 from purple membrane in H2O are compared to those in D2O. It is found that changes in the spectrum occur in the 1,600 - 1,650 cm-1 region as well as in the 800 - 1,000 cm-1 region, but apparently not in the fingerprint region (1,100 - 1,400 cm-1). The possibilities of conformational changes of the retinal chromophore in the light adaptation process as well as the photosynthetic cycle are discussed.

Carbon-13 Chemical Shifts of Retinal Isomers and Their Schiff Bases as Models of Visual Chromophores

Abstract Carbon-13 NMR spectra have been observed for 11- and 13-cis-retinals and their Schiff base linkage compounds with butylamine as a model compound of visual chromophore. The chemical shift changes on going from retinal to Schiff base. The changes in chemical shifts of polyene carbons were discussed in correlation with the changes of π-electron densities.

Specificity of the retinal binding site of bacteriorhodopsin: chemical and stereochemical requirements for the binding of retinol and retinal.

The complexes formed from bacteriopsin and various retinyl compounds were analyzed by fluorescence and absorption spectroscopy. The binding of retinol occurs in two steps. In the first reaction the molecule is fixed in the retinal binding site of the protein. In this state, energy transfer from aromatic amino acid residues to the retinyl moiety is observed. all-trans-Retinal and the 13-, 11-, and 9-cis-retinols are bound in the chromophoric site. In the second reaction the cyclohexene ring and the side chain of the retinyl moiety are forced into a planar conformation. This reaction is mediated by a base (B1) with a pK of 3.8 and requires the oxygen atom but not the free hydroxyl group of retinol, indicating interaction with a group AH (pK greater than or equal to 10.5). The ring-chain planarization reaction is blocked for the 9-cis isomer of retinol. Binding studies with bacterioopsin and retinal isomers reveal that, as in the case of the corresponding retinols, B1 mediates ring-chain planarization in the case of the all-trans, 13-cis, and 11-cis isomers but not with the 9-cis isomer. Reconstitution of the purple complex from the intermediate 430-460-nm chromophore requires the presence of a second base (B2) with a pK of 4.6. This reaction is exclusive for all-trans- and 13-cis-retinal

Symmetric and Asymmetric Bilayers of Retinal

Abstract Asymmetric and symmetric bilayer membrane were formed from monolayers of egg lecithin and various retinal isomers. Capacitance of the asymmetric all-trans retinal/lecithin bilayer is 1.2 ± 0.1 µF/cm2. Asymmetrically formed membranes of 13-cis retinal/lecithin and 9-cis retinal lecithin were found to be 0.83 ± 0.06 and 0.56 ± 0.05 µF/cm2, respectively. Formation of bilayers of the various cis isomers appears to require the presence of solvent. Irradiation of the various retinal/lecithin films, under the proper conditions, leads to bilayers with marked changes in capacitance and conductance.