9-cis-retinal and 13-cis-retinal

Complete and unambiguous (1)H and (13)C NMR chemical shift assignments for all-trans-retinal, 13-cis-retinal, 11-cis-retinal and 9-cis-retinal (1-4) have been established by means of two-dimensional COSY, HSQC, HMBC and NOESY spectroscopic experiments.

Topics: Diterpenes; Magnetic Resonance Spectroscopy; Molecular Structure; Retinaldehyde

We report the use of microcantilevers (MCs) for the detection of three retinoid isomers: 9-cis-retinal, 13-cis-retinal and all-trans-retinal. Detection of synthetic and natural retinoids in topical cosmetic products is important, and their presence can be used to predict reactions with the skin surface. In this study the MC surfaces were functionalized in order to promote the formation of covalent bonds with the chromophores. The lowest mass shift we detected with the functionalized MCs was 1.2 ppt, which is in the range needed by the cosmetics industry. Our results indicate that properly designed and functionalized microcantilevers can be used to construct economical, fast, and sensitive sensors for quality control in cosmetics.

Topics: Biosensing Techniques; Cosmetics; Diterpenes; Isomerism; Retinaldehyde; Surface Properties

RPE65 is essential for the generation of 11-cis retinal. Rod photoreceptors in the RPE65-knockout (Rpe65(-/-)) mouse are known to degenerate slowly with age. This study was designed to examine cone photoreceptors and the expression of cone-specific genes in the Rpe65(-/-) mouse.. Gene expression changes were identified by microarray and confirmed by real-time RT-PCR. Cone photoreceptors were stained by peanut agglutinin (PNA) lectin in the flatmounted retina. The 9- or 11-cis retinal was supplied by intraperitoneal injections.. The short-wavelength (SWL) cone opsin mRNA was markedly decreased at 2 weeks of age, whereas the decrease in the middle-wavelength (MWL) cone opsin mRNA occurred relatively later in age. In contrast, the rhodopsin mRNA level did not show any significant change at all the ages analyzed. Consistent with the cone opsin changes, the cone transducin alpha-subunit mRNA decreased at both 4 and 8 weeks of age, whereas again the rod transducin alpha-subunit did not show any significant change. Rpe65(-/-) mice showed significant cone loss in both the central and ventral retina between 2 and 3 weeks of age. Administration of 9- or 11-cis retinal to Rpe65(-/-) mice 2 weeks of age increased cone density by twofold in these areas.. In the Rpe65(-/-) mouse, the expression of cone-specific genes is downregulated and is accompanied by cone degeneration at early ages. Early administration of 9- or 11-cis retinal can partially prevent cone loss, suggesting that the absence of 11-cis chromophore may be responsible for the early cone degeneration.

Topics: Animals; Carrier Proteins; Cell Count; cis-trans-Isomerases; Diterpenes; Down-Regulation; Eye Proteins; Gene Expression; Gene Expression Profiling; GTP-Binding Protein alpha Subunits; Heterotrimeric GTP-Binding Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Oligonucleotide Array Sequence Analysis; Proteins; Retinal Cone Photoreceptor Cells; Retinal Degeneration; Retinaldehyde; Reverse Transcriptase Polymerase Chain Reaction; Rhodopsin; RNA, Messenger; Rod Opsins; Transducin

Retinal dehydrogenases (RALDHs) convert retinal into retinoic acids (RAs), which are important signaling molecules in embryogenesis and tissue differentiation. We expressed mouse RALDH type 1 (mRALDH1) in Escherichia coli and studied the kinetic properties of the recombinant enzyme for retinal substrates. Purified recombinant mRALDH1 catalyzed the oxidation of all-trans and 9-cis retinal but not 13-cis retinal, and exhibited two pH optimums, 7.8 and 9.4, for all-trans and 9-cis retinal substrates, respectively. The K(m) for all-trans retinal (11.6 micro M) was 3-fold higher than for 9-cis retinal (3.59 micro M). However, the conversion efficiencies of either all-trans or 9-cis retinal to the respective RAs were similar. MgCl(2) inhibited the oxidation of both all-trans and 9-cis retinal. Chloral hydrate and acetaldehyde competitively suppressed all-trans retinal oxidation with inhibition constants (K(i)) of 4.99 and 49.4 micro M, respectively. Retinol, on the other hand, blocked the reaction uncompetitively. These data extend the kinetic characterization of mRALDH1, provide insight into the possible role of this enzyme in the biogenesis of RAs, and should give useful information on the determination of amino acid residues that play crucial roles in the catalysis of all-trans and 9-cis retinal.

Topics: Aldehyde Oxidoreductases; Animals; Diterpenes; Humans; Hydrogen-Ion Concentration; Kinetics; Mice; Rats; Recombinant Proteins; Retinal Dehydrogenase; Retinaldehyde; RNA, Messenger; Species Specificity

Retinal dehydrogenase (RALDH) isozymes catalyze the terminal oxidation of retinol into retinoic acid (RA) that is essential for embryogenesis and tissue differentiation. To understand the role of mouse type 2 RALDH in synthesizing the ligands (all-trans and 9-cis RA) needed to bind and activate nuclear RA receptors, we determined the detailed kinetic properties of RALDH2 for various retinal substrates. Purified recombinant RALDH2 showed a pH optimum of 9.0 for all-trans retinal oxidation. The activity of the enzyme was lower at 37 degrees C compared to 25 degrees C. The efficiency of conversion of all-trans retinal to RA was 2- and 5-fold higher than 13-cis and 9-cis retinal, respectively. The K(m) for all-trans and 13-cis retinal were similar (0.66 and 0.62 microM, respectively). However, the K(m) of RALDH2 for 9-cis retinal substrate (2.25 microM) was 3-fold higher compared to all-trans and 13-cis retinal substrates. Among several reagents tested for their ability to either inhibit or activate RALDH2, citral and para-hydroxymercuribenzoic acid (p-HMB) inhibited and MgCl(2) activated the reaction. Comparison of the kinetic properties of RALDH2 for retinal substrates and its activity towards various reagents with those of previously reported rat kidney RALDH1 and human liver aldehyde dehydrogenase-1 showed distinct differences. Since RALDH2 has low K(m) and high catalytic efficiency for all-trans retinal, it may likely be involved in the production of all-trans RA in vivo.

Topics: Aldehyde Oxidoreductases; Alitretinoin; Animals; Catalysis; Cloning, Molecular; Diterpenes; Gene Expression Regulation, Enzymologic; Hydrogen-Ion Concentration; Isotretinoin; Kinetics; Mice; Recombinant Proteins; Retinal Dehydrogenase; Retinaldehyde; Temperature; Tretinoin; Vitamin A

Photoisomerization of 11-cis-retinal to all-trans-retinal triggers phototransduction in the retinal photoreceptor cells and causes ultimately the sensation of vision. 11-cis-Retinal is enzymatically regenerated through a complex set of reactions in adjacent retinal pigment epithelial cells (RPE). In this study using all-trans-9-desmethylretinol (lacking the C(19) methyl group) and all-trans-13-desmethylretinol (lacking the C(20) methyl group), we explored the effects of C(19) and C(20) methyl group removals on isomerization of these retinols in RPE microsomes. The C(19) methyl group may be involved in the substrate activation, whereas the C(20) methyl group causes steric hindrance with a proton in position C(10) of 11-cis-retinol; thus, removal of this group could accelerate isomerization. We found that all-trans-9-desmethylretinol and all-trans-13-desmethylretinol are isomerized to their corresponding 11-cis-alcohols, although with lower efficiencies than isomerization of all-trans-retinol to 11-cis-retinol. These findings make the mechanism of isomerization through the C(19) methyl group unlikely, because in the case of 9-desmethylretinol, the isomerization would have to progress by proton abstraction from electron-rich olefinic C(9). The differences between all-trans-retinol, all-trans-9-desmethylretinol, and all-trans-13-desmethylretinol appear to be a consequence of the enzymatic properties, and binding affinities of the isomerization system, rather than differences in the chemical or thermodynamic properties of these compounds. This observation is also supported by quantum chemical calculations. It appears that both methyl groups are not essential for the isomerization reaction and are not likely involved in formation of a transition stage during the isomerization process.

Topics: Alcohol Oxidoreductases; Animals; Apoproteins; Carrier Proteins; Cattle; Diterpenes; Isomerism; Pigment Epithelium of Eye; Protein Conformation; Quantum Theory; Retinaldehyde; Retinoids; Serum Albumin, Bovine

Protonation changes of the protein occur during the reconstitution of bacteriorhodopsin from bacterio-opsin and all-trans retinal in the purple membrane of Halobacterium halobium. The protonation changes are conveniently determined from measures of the pH changes after photoisomerisation of 9-cis retinal in apomembrane preparations, which induces the reconstitution. In addition, to the omega-amino group of the lysine which is involved in the condensation of retinal and bacterio-opsin, the dissociation equilibria of at least two other amino acid residues are changed during the reconstitution. The results are consistent with a proposed model of chromophore structure in which an interaction of the Schiff's base occurs with two protonable amino acid residues.

Topics: Apoproteins; Bacteriorhodopsins; Carotenoids; Chemical Phenomena; Chemistry; Diterpenes; Halobacterium; Hydrogen-Ion Concentration; Isomerism; Light; Models, Chemical; Protons; Retinaldehyde; Ultraviolet Rays; Vitamin A