X-ray crystallographic analysis of 9-cis-rhodopsin, a model analogue visual pigment

Recent progress in high-resolution structural study of rhodopsin has been enabled by a novel selective extraction procedure with rod photoreceptor cells. In this study, we applied the method for rapid and efficient preparation of a purified analogue pigment using bovine rod outer segment membranes with 9-cis-retinal. After complete bleaching of the membranes and subsequent regeneration with the exogenous retinal, 9-cis-rhodopsin is selectively extracted from the membranes using combination of zinc and heptylthioglucoside. The solubilized sample, even with a small amount of contaminating retinal oximes, is shown to be pure enough for three-dimensional crystallization. The X-ray diffraction from 9-cis-rhodopsin crystals was examined and the electron density map at 2.9 angstroms resolution in the chromophore region can be fitted well with the model of 9-cis-retinal Schiff base.     

Microtubules and actin filaments in teleost visual cone elongation and contraction.

Teleost retinal cones contract in light and elongate in darkness. This paper describes the disposition of microtubules and cytoplasmic filaments in cone cells of 2 species of fish (Haemulon sciurus and Lutjanus griseus). In Haemulon, the neck-like "myoid" region of the cone changes in length from 5 mu to 75 mu. Maximal observed rates of elongation and contraction are comparable to that of chromosome movement in mitosis (2-3 mu/min). Microtubules presumably participate in cone elongation, since numerous longitudinal microtubules are present in the myoid region, and colchicine blocks dark-induced elongation. Myoid shortening, on the other hand, appears to be an active contractile process. Disruption of microtubules in dark-adapted cones does not produce myoid shortening in the absence of light, and light-induced myoid shortening is blocked by cytochalasin-B. Cone cells possess longitudinally-oriented thin filaments which bind myosin subfragment-1 to form arrowhead complexes typical of muscle actin. Myoid thin filaments are clearly observed in negatively stained preparations of isolated cones which have been disrupted with detergent after attachment to grids. These myoid filaments are not, however, generally preserved by conventional fixation, though bundles of thin filaments are preserved in other regions of the cell. Thus, actin filaments are poorly retained by fixation in precisely the region of the cone cell where contraction occurs. Cone cells also possess longitudinally-oriented thick filaments 130-160 A in diameter. That these thick filaments may be myosin is suggested by the presence of side-arms with approximately 150 A periodicity. The linear organization of the contractile apparatus of the retinal cone cell makes this cell a promising model for morphological characterization of the disposition of actin and myosin filaments during contraction in a nonmuscle cell.     

Convergent stereoselective synthesis of the visual pigment A2E

[chemical reaction: see text]. A stereoselective total synthesis of the visual pigment A2E has been achieved with use of palladium-catalyzed cross-coupling reactions in all key steps: a regioselective Suzuki or Negishi coupling of 2,4-dibromopyridine, a Sonogashira reaction, and a double Stille cross-coupling to complete the bispolyenyl skeleton.     

Wavelength regulation in visual pigment chromophore. Large induced bathochromic shifts in retinol and related polyenes

Abstract— Low temperature techniques are used to record spectra of long wavelength absorbing electrochromic species formed from retinol and related compounds. Formation of these electrochromic species is induced either by iodine or trichloroacetic acid which merely act as acids. The electrochromic species are the corresponding carbonium ions of the respective polyenes. The resulting carbonium ions of a given polyene are identical regardless of whether they are formed by iodine or trichloroacetic acid. The molecular structures of the carbonium ions can be written with certainty. Consequently, the exact nature of the π-electron system in the polyene and its carbonium ion may be elucidated.     

Molecular mechanics of retinal: Twisting mechanism of chromophore in visual pigment

In a model calculation pulli 11-cis retinal (9-cis retinal) from both end sides, the 11–12 double bond (9–10 double bond) is found to be selectively twisted. This property is promising for the twisting mechanism of retinal chromophore in visual pigment as assumed by the Kakitani and Kakitani torsion model.     

On the nature of excited electronic states in cyanine dyes: implications for visual pigment spectra

CNDO/S CI calculations are carried out on polyenes and on cyanine dyes. In contrast to polyenes, doubly excited configurations have a strong effect on the first optically allowed excited state in cyanines. Protonated Schiff bases of retinal are closely related to cyanine dyes, with important consequences for models of visual pigment spectra and photochemistry.     

Carbon-13 NMR spectra of all-trans-N-retinylidene-n-butylamine as an analogue of the Schiff's base linkage-compound in visual pigment

Carbon-13 NMR spectra have been obtained for all-trans-N-retinylidene-n-butylamine as an analogue of the Schiff's base linkage compound in visual pigment. The chemical shift changes on going from all-trans-retinal₁ to all-trans-N-retinylidene-n-butylamine are observed. These changes indicate the collapse of the bond alternation in conjugated polyene carbons, whereas N-protonation in all-trans-N-retinylidene-n-butylamine does not affect the bond alternation.     

Metabolites of pyrenomycetes XI. Structure of Aurocitrin,a new antibacterial pigment from Hypocrea citrina.

Isolation and structure elucidation of aurocitrin, a new antibiotic metabolite from two varieties of the hypocrealean fungus, H. citrina, is reported.     

Photodecomposition of Pigment Yellow 74, a pigment used in tattoo inks

Tattooing has become a popular recreational practice among younger adults over the past decade. Although some of the pigments used in tattooing have been described, very little is known concerning the toxicology, phototoxicology or photochemistry of these pigments. Seven yellow tattoo inks were obtained from commercial sources and their pigments extracted, identified and quantitatively analyzed. The monoazo compound Pigment Yellow 74 (PY74; CI 11741) was found to be the major pigment in several of the tattoo inks. Solutions of commercial PY74 in tetrahydrofuran (THF) were deoxygenated using argon gas, and the photochemical reaction products were determined after exposure to simulated solar light generated by a filtered 6.5 kW xenon arc lamp. Spectrophotometric and high-pressure liquid chromatography (HPLC) analyses indicated that PY74 photodecomposed to multiple products that were isolated using a combination of silica chromatography and reversed-phase HPLC. Three of the major photodecomposition products were identified by nuclear magnetic resonance and mass spectrometry as N-(2-methoxyphenyl)-3-oxobutanamide (o-acetoacetanisidide), 2-(hydroxyimine)-N-(2-methoxyphenyl)-3-oxobutanamide and N,N'-bis(2-methoxyphenyl)urea. These results demonstrate that PY74 is not photostable in THF and that photochemical lysis occurs at several sites in PY74 including the hydrazone and amide groups. The data also suggest that the use of PY74 in tattoo inks could potentially result in the formation of photolysis products, resulting in toxicity at the tattoo site after irradiation with sunlight or more intense light sources.     

Automated, fluorescence-based approach for the specific diagnosis of chicken coccidiosis.

We have established a fluorescence-based electrophoretic approach for the specific identification of all seven currently recognised species of Eimeria infecting chickens. The second internal transcribed spacer (ITS-2) of ribosomal DNA is amplified by polymerase chain reaction (PCR) from any of the seven species using a single set of oligonucleotide primers (of which the reverse one is fluorescently labelled). The amplicons are heat-denatured, subjected to denaturing polyacrylamide gel electrophoresis in a 377 DNA sequencer (ABI). The chromatograms produced are stored electronically and then analysed using GeneScan 3.1 software. Using control DNA samples representing monospecific lines of Eimeria, regions in the chromatograms have been defined for the specific identification of each of the seven species, although some variation in the chromatograms (reflecting population variation) was detectable within two species. Electrophoretic reading and analysis is carried out automatically using a computer imaging system, thus making it a time- and cost-effective approach. It is well suited for high-throughput diagnostic screening of oocyst samples and should find applicability as a tool for prevalence studies, monitoring of coccidiosis outbreaks and the quality control of vaccines.     

Katorazone,a new yellow pigment with a 2-azaquinone-phenylhydrazone structure produced by Streptomyces sp. IFM 11299

An unusual alkaloid with a 2-azaquinone-phenylhydrazone structure, katorazone (1), and other metabolites were isolated from the ethyl acetate extract of Streptomyces sp. IFM 11299. The chemical structure of katorazone (1) was elucidated by 1D and 2D NMR analyses together with HR-ESI mass spectrometry. Katorazone (1) showed a synergistic effect in combination with TRAIL and decreased the viability of human gastric adenocarcinoma (AGS) cells.     

Hinnuliquinone,a bis-indolyl-2,5-dihydroxybenzoquinone pigment from nodulisphorium hinnuleum

A pigment, hinnuliquinone, isolated from the fungus $\text{Nodulisporium}$$\text{hinnuleum}$, has been shown to be 2,5-bis-[2-(1,1,-dimethyl-2-propenyl)1H-indol-3-yl-]-3,6-dihydroxy-2,5-cyclohexadiene-1,4-dione and to be biosynthesized from tryptophan and mevalonic acid.     

Phototoxicity in human retinal pigment epithelial cells promoted by hypericin, a component of St. John's wort

St. John's wort (SJW), an over-the-counter antidepressant, contains hypericin, which absorbs light in the UV and visible ranges. In vivo studies have determined that hypericin is phototoxic to skin and our previous in vitro studies with lens tissues have determined that it is potentially phototoxic to the human lens. To determine if hypericin might also be phototoxic to the human retina, we exposed human retinal pigment epithelial (hRPE) cells to 10(-7) to 10(-5) M hypericin. Fluorescence emission detected from the cells (lambda(ex) = 488 nm; lambda(em) = 505 nm) confirmed hypericin uptake by human RPE. Neither hypericin exposure alone nor visible light exposure alone reduced cell viability. However when irradiated with 0.7 J cm(-2) of visible light (lambda > 400 nm) there was loss of cell viability as measured by MTS and lactate dehydrogenase assays. The presence of hypericin in irradiated hRPE cells significantly changed the redox equilibrium of glutathione and a decrease in the activity of glutathione reductase. Increased lipid peroxidation as measured by the thiobarbituric acid reactive substances assay correlated to hypericin concentration in hRPE cells and visible light radiation. Thus, ingested SJW is potentially phototoxic to the retina and could contribute to retinal or early macular degeneration.     

Sensitivity of the retina to radiation damage as a function of wavelength.

Abstract—Exposure of the retina of the rhesus monkey to visible and infrared radiation from CW optical sources like the Sun, xenon lamps, etc. produces small lesions or scotomata which may be classified as thermal or photochemical, depending on the wavelength and duration of exposure. The action spectrum for the production of retinal lesions has been determined for eight monochromatic laser wavelengths extending from 1064 to 441 nm. The corneal power required to produce a lesion decreases by three orders of magnitude in going from 1064 to 441 nm. Exposure to 1064 nm radiation for 1000 s produces a typical thermal lesion at elevated retinal temperatures. whereas a 1000 s exposure to 441 nm light produces a photochemical lesion at power levels too low to raise the retinal temperature by an appreciable amount (<0.1°). The two types of lesion have entirely different characteristics as will be discussed in some detail. The photopathology of thc photochemical lesion has been studied at postexposure times ranging from 1 h to 90 days and will be demonstrated in a number of histological slides. Moreover, this photopathology correlates well with monocular visual acuity tests in the rhesus monkey as defined by the Landolt ring technique. To further elucidate the differential effects on the retina of short vs long wavelength CW radiation, we have divided a simulated solar spectrum at sea level into two spectral bands. 400–800 nm and 700–1400 nm, and determined the radiant exposures required to produce very mild lcsions on the rhesus retina for exposure times of 1, 10, 100 and 1000 s. To correlate our data with solar retinitis and eclipse blindness the image diameter or spot size on the retina was 159 μm, corresponding to the image size of the Sun on the human retina. Exposure to the 400–800 nm spectrum for durations of 10 s or greater required approximately 400 J/cm² to produce a mild photochemical lesion. Reciprocity is maintained over the exposure range 10–1000 s. Radiant exposure to the 700–1400 nm spectrum, on the other hand, required roughly 69,100 J/cm² for a 1000 s exposure. This was a mild thermal lesion. We were unable to produce a lesion for exposure times less than 1000 s. We interpret these data to mean that solar retinitis and eclipse blindness are primarily photochemical events produced by the short wavelength component of the solar spectrum, and that the infrared component of the solar spectrum plays only a minor role in these retinal pathologies.     

Biomimetic synthesis of carthamin,a red pigment in safflower petals,via oxidative decarboxylation

Carthamin, a natural red pigment derived from safflower, was discovered centuries ago and has been utilized in several industries such as dye, food coloring, and natural medicine industries. Although it has been used since Egyptian times, its structural features were only determined in the mid-1900s when pioneering studies were conducted to elucidate the carthamin structure and assign its stereochemistry. Reproducing the complex biosynthesis of carthamin has been a common research goal since then. However, this has been rendered complex because of the unstable precursors and low yields from the multiple steps involved in the synthesis. Herein, we report the total synthesis of carthamin via a four-step procedure using (S)-C-β-D-glucosylquinochalcone as the starting compound. The synthesis involves mild conditions, short reaction time, and readily available compounds. After de-O-methylation, condensation, oxidative decarboxylation, and saponification, carthamin was obtained in an overall yield of 9.5% through the proposed method.