retinaldehyde and 13-cis-retinal
retinaldehyde has been researched along with 13-cis-retinal* in 29 studies
Reviews
1 review(s) available for retinaldehyde and 13-cis-retinal
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Chemical dynamics in proteins: the photoisomerization of retinal in bacteriorhodopsin.
Chemical dynamics in proteins are discussed, with bacteriorhodopsin serving as a model system. Ultrafast time-resolved methods used to probe the chemical dynamics of retinal photoisomerization in bacteriorhodopsin are discussed, along with future prospects for ultrafast time-resolved crystallography. The photoisomerization of retinal in bacteriorhodopsin is far more selective and efficient than in solution, the origins of which are discussed in the context of a three-state model for the photoisomerization reaction coordinate. The chemical dynamics are complex, with the excited-state relaxation exhibiting a multiexponential decay with well-defined rate constants. Possible origins for the two major components are also discussed. Topics: Bacteriorhodopsins; Diterpenes; Halobacterium salinarum; Isomerism; Kinetics; Light; Models, Chemical; Protein Conformation; Retinaldehyde; Spectrum Analysis; Thermodynamics | 1998 |
Trials
1 trial(s) available for retinaldehyde and 13-cis-retinal
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Temozolomide and 13-cis retinoic acid in patients with anaplastic gliomas: a prospective single-arm monocentric phase-II study (RNOP-05).
The objective of this prospective, monocentric phase-II pilot study was to evaluate toxicity and efficacy of neoadjuvant temozolomide (TMZ) and 13-cis retinoic acid (13-cRA) treatment in patients with newly diagnosed anaplastic gliomas after total or subtotal tumor resection. The primary endpoint of the study was median progression-free survival (PFS). Secondary endpoints were toxicity and PFS rates at 6, 12 and 24 months. Thirty-two adult patients were included in the study and treated with a median number of 10 TMZ and 13-cRA cycles (range 1-26). The majority of patients had favorable prognostic factors characterized by young age, complete resection, oligodendroglial histology, 1p/19q co-deletion, O6-methylguanine-DNA methyltransferase (MGMT) promotor methylation and isocitrate dehydrogenase 1 (IDH1) mutation. Grade 3/4 myelotoxicity occurred in 5/32 patients, and about 90% of patients suffered from grade 2/3 adverse events attributable to 13-cRA. The median PFS was 37.8 months (95% CI 22.2-53.4). The 6-, 12- and 24-month PFS rates were 84.4, 75 and 42.4%. The extent of tumor resection was the only prognostic factor associated with better PFS. TMZ and 13-cRA treatment did not improve PFS when retrospectively compared to the TMZ-treated group within the randomized NOA-04 phase-III trial. In conclusion, 13-cRA addition to TMZ in a neoadjuvant setting showed acceptable toxicity, but did not yield an advantage in PFS in patients with newly diagnosed anaplastic gliomas after total or subtotal tumor resection. Topics: Administration, Oral; Adolescent; Adult; Antineoplastic Combined Chemotherapy Protocols; Brain Neoplasms; Dacarbazine; Disease-Free Survival; Diterpenes; DNA Modification Methylases; DNA Repair Enzymes; Drug Administration Schedule; Female; Glioma; Humans; Isocitrate Dehydrogenase; Male; Middle Aged; Mutation; Prospective Studies; Retinaldehyde; Temozolomide; Tumor Suppressor Proteins; Young Adult | 2011 |
Other Studies
27 other study(ies) available for retinaldehyde and 13-cis-retinal
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Low-temperature Raman spectroscopy of sodium-pump rhodopsin from
We carried out the low-temperature Raman measurement of a sodium pump rhodopsin from Indibacter alkaliphilus (IaNaR) and examined the primary structural change for the light-driven Na+ pump. We observed that photoexcitation of IaNaR produced the distorted 13-cis retinal chromophore in the presence of Na+, while the structural distortion was significantly relaxed in the absence of Na+. This structural difference of the chromophore with/without Na+ was attributed to the Na+ binding to the protein, which alters the active site. Using the spectral sensitivity to the ion binding, we found that IaNaR had a second Na+ binding site in addition to the one already specified on the extracellular surface. To date, the Na+ binding has not been considered as a prerequisite for Na+ transport. However, this study provides insight that the protein structural change induced by the ion binding involved the formation of an R108-D250 salt bridge, which has critical importance in the active transport of Na+. Topics: Bacterial Proteins; Bacteroidetes; Biological Transport, Active; Catalytic Domain; Cation Transport Proteins; Cold Temperature; Crystallography, X-Ray; Diterpenes; Molecular Conformation; Mutation; Retinaldehyde; Rhodopsins, Microbial; Sodium; Spectrum Analysis, Raman | 2021 |
The Desensitized Channelrhodopsin-2 Photointermediate Contains 13 -cis, 15 -syn Retinal Schiff Base.
Channelrhodopsin-2 (ChR2) is a light-gated cation channel and was used to lay the foundations of optogenetics. Its dark state X-ray structure has been determined in 2017 for the wild-type, which is the prototype for all other ChR variants. However, the mechanistic understanding of the channel function is still incomplete in terms of structural changes after photon absorption by the retinal chromophore and in the framework of functional models. Hence, detailed information needs to be collected on the dark state as well as on the different photointermediates. For ChR2 detailed knowledge on the chromophore configuration in the different states is still missing and a consensus has not been achieved. Using DNP-enhanced solid-state MAS NMR spectroscopy on proteoliposome samples, we unambiguously determined the chromophore configuration in the desensitized state, and we show that this state occurs towards the end of the photocycle. Topics: Cations; Channelrhodopsins; Chlamydomonas reinhardtii; Diterpenes; Light; Magnetic Resonance Spectroscopy; Photochemical Processes; Photons; Protein Conformation; Retinaldehyde; Schiff Bases | 2021 |
Formation Mechanism of Ion Channel in Channelrhodopsin-2: Molecular Dynamics Simulation and Steering Molecular Dynamics Simulations.
Channelrhodopsin-2 (ChR2) is a light-activated and non-selective cationic channel protein that can be easily expressed in specific neurons to control neuronal activity by light. Although ChR2 has been extensively used as an optogenetic tool in neuroscience research, the molecular mechanism of cation channel formation following retinal photoisomerization in ChR2 is not well understood. In this paper, studies of the closed and opened state ChR2 structures are presented. The formation of the cationic channel is elucidated in atomic detail using molecular dynamics simulations on the all-trans-retinal (ChR2-trans) configuration of ChR2 and its isomerization products, 13-cis-retinal (ChR2-cis) configuration, respectively. Photoisomerization of the retinal-chromophore causes the destruction of interactions among the crucial residues (e.g., E90, E82, N258, and R268) around the channel and the extended H-bond network mediated by numerous water molecules, which opens the pore. Steering molecular dynamics (SMD) simulations show that the electrostatic interactions at the binding sites in intracellular gate (ICG) and central gate (CG) can influence the transmembrane transport of Na Topics: Channelrhodopsins; Chlamydomonas reinhardtii; Diterpenes; Ion Transport; Isomerism; Molecular Dynamics Simulation; Plant Proteins; Protein Conformation; Protein Multimerization; Retinaldehyde | 2019 |
Mapping the ultrafast vibrational dynamics of all-trans and 13-cis retinal isomerization in Anabaena Sensory Rhodopsin.
Discrepancies in the isomerization dynamics and quantum yields of the trans and cis retinal protonated Schiff base is a well-known issue in the context of retinal photochemistry. Anabaena Sensory Rhodopsin (ASR) is a microbial retinal protein that comprises a retinal chromophore in two ground state (GS) conformations: all-trans, 15-anti (AT) and 13-cis, 15-syn (13C). In this study, we applied impulsive vibrational spectroscopic techniques (DFWM, pump-DFWM and pump-IVS) to ASR to shed more light on how the structural changes take place in the excited state within the same protein environment. Our findings point to distinct features in the ground state structural conformations as well as to drastically different evolutions in the excited state manifold. The ground state vibrational spectra show stronger Raman activity of the C14-H out-of-plane wag (at about 805 cm-1) for the 13C isomer than that for the AT isomer, which hints at a pre-distortion of 13C in the ground state. Evolution of the Raman frequency after interaction with the actinic pulse shows a blue-shift for the C[double bond, length as m-dash]C stretching and CH3 rocking mode for both isomers. For AT, however, the blue-shift is not instantaneous as observed for the 13C isomer, rather it takes more than 200 fs to reach the maximum frequency shift. This frequency blue-shift is rationalized by a decrease in the effective conjugation length during the isomerization reaction, which further confirms a slower formation of the twisted state for the AT isomer and corroborates the presence of a barrier in the excited state trajectory previously predicted by quantum chemical calculations. Topics: Anabaena; Bacterial Proteins; Diterpenes; Retinaldehyde; Sensory Rhodopsins; Stereoisomerism; Vibration | 2018 |
Pre-gating conformational changes in the ChETA variant of channelrhodopsin-2 monitored by nanosecond IR spectroscopy.
Light-gated ion permeation by channelrhodopsin-2 (ChR2) relies on the photoisomerization of the retinal chromophore and the subsequent photocycle, leading to the formation (on-gating) and decay (off-gating) of the conductive state. Here, we have analyzed the photocycle of a fast-cycling ChR2 variant (E123T mutation, also known as ChETA), by time-resolved UV/vis, step-scan FT-IR, and tunable quantum cascade laser IR spectroscopies with nanosecond resolution. Pre-gating conformational changes rise with a half-life of 200 ns, silent to UV/vis but detected by IR spectroscopy. They involve changes in the peptide backbone and in the H-bond of the side chain of the critical residue D156. Thus, the P1(500) intermediate must be separated into early and late states. Light-adapted ChR2 contains a mixture of all-trans and 13-cis retinal in a 70:30 ratio which are both photoactive. Analysis of ethylenic and fingerprint vibrations of retinal provides evidence that the 13-cis photocycle recovers in 1 ms. This recovery is faster than channel off-gating and most of the proton transfer reactions, implying that the 13-cis photocycle is of minor functional relevance for ChR2. Topics: Darkness; Diterpenes; Kinetics; Mutation; Photolysis; Protein Conformation; Retinaldehyde; Rhodopsin; Spectroscopy, Fourier Transform Infrared; Static Electricity; Time Factors; Vibration | 2015 |
Molecular Dynamics of Channelrhodopsin at the Early Stages of Channel Opening.
Channelrhodopsin (ChR) is a light-gated cation channel that responds to blue light. Since ChR can be readily expressed in specific neurons to precisely control their activities by light, it has become a powerful tool in neuroscience. Although the recently solved crystal structure of a chimeric ChR, C1C2, provided the structural basis for ChR, our understanding of the molecular mechanism of ChR still remains limited. Here we performed electrophysiological analyses and all-atom molecular dynamics (MD) simulations, to investigate the importance of the intracellular and central constrictions of the ion conducting pore observed in the crystal structure of C1C2. Our electrophysiological analysis revealed that two glutamate residues, Glu122 and Glu129, in the intracellular and central constrictions, respectively, should be deprotonated in the photocycle. The simulation results suggested that the deprotonation of Glu129 in the central constriction leads to ion leakage in the ground state, and implied that the protonation of Glu129 is important for preventing ion leakage in the ground state. Moreover, we modeled the 13-cis retinal bound; i.e., activated C1C2, and performed MD simulations to investigate the conformational changes in the early stage of the photocycle. Our simulations suggested that retinal photoisomerization induces the conformational change toward channel opening, including the movements of TM6, TM7 and TM2. These insights into the dynamics of the ground states and the early photocycle stages enhance our understanding of the channel function of ChR. Topics: Bacteriorhodopsins; Crystallography, X-Ray; Diterpenes; Electrophysiology; Glutamine; HEK293 Cells; Humans; Ion Channel Gating; Models, Molecular; Molecular Dynamics Simulation; Protein Interaction Domains and Motifs; Retinaldehyde; Rhodopsin | 2015 |
Light-Dark Adaptation of Channelrhodopsin Involves Photoconversion between the all-trans and 13-cis Retinal Isomers.
Channelrhodopsins (ChR) are light-gated ion channels of green algae that are widely used to probe the function of neuronal cells with light. Most ChRs show a substantial reduction in photocurrents during illumination, a process named "light adaptation". The main objective of this spectroscopic study was to elucidate the molecular processes associated with light-dark adaptation. Here we show by liquid and solid-state nuclear magnetic resonance spectroscopy that the retinal chromophore of fully dark-adapted ChR is exclusively in an all-trans configuration. Resonance Raman (RR) spectroscopy, however, revealed that already low light intensities establish a photostationary equilibrium between all-trans,15-anti and 13-cis,15-syn configurations at a ratio of 3:1. The underlying photoreactions involve simultaneous isomerization of the C(13)═C(14) and C(15)═N bonds. Both isomers of this DAapp state may run through photoinduced reaction cycles initiated by photoisomerization of only the C(13)═C(14) bond. RR spectroscopic experiments further demonstrated that photoinduced conversion of the apparent dark-adapted (DAapp) state to the photocycle intermediates P500 and P390 is distinctly more efficient for the all-trans isomer than for the 13-cis isomer, possibly because of different chromophore-water interactions. Our data demonstrating two complementary photocycles of the DAapp isomers are fully consistent with the existence of two conducting states that vary in quantitative relation during light-dark adaptation, as suggested previously by electrical measurements. Topics: Animals; Channelrhodopsins; Dark Adaptation; Diterpenes; Insecta; Isomerism; Photic Stimulation; Pichia; Retinaldehyde | 2015 |
Complete NMR assignment of retinal and its related compounds.
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 | 2013 |
Ultrafast photochemistry of light-adapted and dark-adapted bacteriorhodopsin: effects of the initial retinal configuration.
Femtosecond spectroscopy is used to compare photochemical dynamics in light-adapted and dark-adapted bacteriorhodopsin (BR). The retinal prosthetic group is initially all-trans in the former, while it is nearly a 1:1 mixture with 13-cis in the latter. Comparing photochemistry in both serves to assess how the initial retinal configuration influences internal conversion and photoisomerization dynamics. Contrary to an earlier study, our results show that after excitation of the 13-cis form it crosses back to the ground state much more rapidly than the biologically active all-trans reactant. A similar result was recently obtained for another microbial retinal protein, Anabaena Sensory Rhodospin (ASR), which can be toggled by light between two analogous ground state configurations. Together, these studies suggest that this disparity in rates may be a general trend in the photochemistry of microbial retinal proteins. This may bear as well on the well-known enhancement in photoisomerization rates going from microbial retinal proteins to the visual pigments, as the latter also start the course of photoreception in a cis retinal configuration, in that case 11-cis. In lieu of indications for pretwisting or straining of the 13-cis retinal forms of BR and ASR, akin to those reported for rhodopsin, current results challenge many of the mechanisms held responsible for the ballistic photochemical dynamics observed in visual pigment. Topics: Adaptation, Ocular; Bacteriorhodopsins; Dark Adaptation; Diterpenes; Halobacterium salinarum; Isomerism; Photochemical Processes; Retinaldehyde | 2012 |
An amino acid residue (S201) in the retinal binding pocket regulates the photoreaction pathway of phoborhodopsin.
Phoborhodopsin from Halobacterium salinarum (salinarum phoborhodopsin, spR also called HsSR II) is a photoreceptor for the negative phototaxis of the bacterium. A unique feature of spR is the formation of a shorter wavelength photoproduct, P480, observed at liquid nitrogen temperature beside the K intermediate. Formation of similar photoproduct has not been reported in the other microbial rhodopsins. This photoproduct showed its maximum absorbance wavelength (λ(max)) at 482 nm and can thermally revert back to spR above -160 °C. It was revealed that P480 is a photoproduct of K intermediate by combination of an irradiation and warming experiment. Fourier transform infrared (FTIR) difference spectrum of P480 from spR in C-C stretching vibration region showed similar features with that of K intermediate, suggesting that P480 has a 13-cis-retinal chromophore. The appearance of a broad positive band at 1214 cm(-1) in the P480-spR spectrum suggested that configuration around C9═C10 likely be different between P480 and K intermediate. Vibrational bands in HOOP region (1035 to 900 cm(-1)) suggested that the chromophore distortion in K intermediate was largely relaxed in P480. The amount of P480 formed by the irradiation was greatly decreased by amino acid replacement of S201 with T, suggesting S201 was involved in the formation of P480. According to the crystal structure of pharaonis phoborhodopsin (ppR), a homologue of spR found in Natronomonas pharaonis, S201 should locate near the C14 of retinal chromophore. Thus, the interaction between S201 and C14 might be the main factor affecting formation of P480. Topics: Amino Acid Substitution; Amino Acids; Diterpenes; Halorhodopsins; Mutation; Natronobacterium; Photochemistry; Protein Binding; Retinaldehyde; Sensory Rhodopsins; Spectroscopy, Fourier Transform Infrared | 2011 |
Cantilever-based sensor for the detection of different chromophore isomers.
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 | 2007 |
FTIR study of the photoisomerization processes in the 13-cis and all-trans forms of Anabaena sensory rhodopsin at 77 K.
Archaeal-type rhodopsins can accommodate either all-trans- or 13-cis,15-syn-retinal in their chromophore binding site in the dark, but only the former isomer is functionally important. In contrast, Anabaena sensory rhodopsin (ASR), an archaeal-type rhodopsin found in eubacteria, exhibits a photochromic interconversion of both forms, suggesting that ASR functions as a photosensor which interacts with its 14 kDa soluble transducer differently in the all-trans and 13-cis,15-syn forms. In this study, we applied low-temperature Fourier transform infrared (FTIR) spectroscopy to the 13-cis,15-syn form of ASR (13C-ASR) at 77 K and compared the local structure around the chromophore and its structural changes upon retinal photoisomerization with those of the all-trans form (AT-ASR) [Furutani, Y., Kawanabe, A., Jung, K. H., and Kandori, H. (2005) Biochemistry 44, 12287-12296]. By use of [zeta-15N]lysine-labeled ASR, we identified the N-D stretching vibrations of the Schiff base (in D2O) at 2165 cm(-1) for 13C-ASR and at 2163 and 2125 cm(-1) for AT-ASR. The frequencies indicate strong hydrogen bonds of the Schiff base with a water molecule for both 13C-ASR and AT-ASR. In contrast, the N-D stretching vibration appears at 2351 cm(-1) and at 2483 cm(-1) for the K states of 13C-ASR (13C-ASR(K)) and AT-ASR (AT-ASR(K)), respectively, indicating that the Schiff base still forms a hydrogen bond in 13C-ASR(K). Rotational motion of the Schiff base upon retinal isomerization is probably smaller for 13C-ASR than for AT-ASR, the latter altering hydrogen bonding of the Schiff base similar to bacteriorhodopsin (BR), a light-driven proton pump. Appearance of several hydrogen-out-of-plane vibrations and amide I vibrations in 13C-ASR(K), but not in AT-ASR(K), suggests that structural changes are distributed widely along the polyene chain for 13C-ASR. On the other hand, retinal photoisomerization in AT-ASR breaks the hydrogen bond of the Schiff base, and localized structural changes in the Schiff base region are induced. Topics: Anabaena; Cold Temperature; Diterpenes; Isomerism; Retinaldehyde; Sensory Rhodopsins; Spectroscopy, Fourier Transform Infrared | 2006 |
Chemistry. Controlling biological functions.
Topics: Bacteriorhodopsins; Diterpenes; Isomerism; Light; Photochemistry; Quantum Theory; Retinaldehyde | 2006 |
Coherent control of retinal isomerization in bacteriorhodopsin.
Optical control of the primary step of photoisomerization of the retinal molecule in bacteriorhodopsin from the all-trans to the 13-cis state was demonstrated under weak field conditions (where only 1 of 300 retinal molecules absorbs a photon during the excitation cycle) that are relevant to understanding biological processes. By modulating the phases and amplitudes of the spectral components in the photoexcitation pulse, we showed that the absolute quantity of 13-cis retinal formed upon excitation can be enhanced or suppressed by +/-20% of the yield observed using a short transform-limited pulse having the same actinic energy. The shaped pulses were shown to be phase-sensitive at intensities too low to access different higher electronic states, and so these pulses apparently steer the isomerization through constructive and destructive interference effects, a mechanism supported by observed signatures of vibrational coherence. These results show that the wave properties of matter can be observed and even manipulated in a system as large and complex as a protein. Topics: Algorithms; Bacteriorhodopsins; Diterpenes; Halobacterium salinarum; Isomerism; Kinetics; Lasers; Light; Photochemistry; Photons; Quantum Theory; Retinaldehyde; Thermodynamics | 2006 |
Downregulation of cone-specific gene expression and degeneration of cone photoreceptors in the Rpe65-/- mouse at early ages.
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 | 2005 |
Crystal structure of the 13-cis isomer of bacteriorhodopsin in the dark-adapted state.
The atomic structure of the trans isomer of bacteriorhodopsin was determined previously by using a 3D crystal belonging to the space group P622. Here, a structure is reported for another isomer with the 13-cis, 15-syn retinal in a dark-adapted crystal. Structural comparison of the two isomers indicates that retinal isomerization around the C13[double bond]C14 and the C15[double bond]N bonds is accompanied by noticeable displacements of a few residues in the vicinity of the retinal Schiff base and small re-arrangement of the hydrogen-bonding network in the proton release channel. On the other hand, aromatic residues surrounding the retinal polyene chain were found to scarcely move during the dark/light adaptation. This result suggests that variation in the structural rigidity within the retinal-binding pocket is one of the important factors ensuring the stereospecific isomerization of retinal. Topics: Bacteriorhodopsins; Crystallography, X-Ray; Darkness; Diterpenes; Halobacterium salinarum; Hydrogen Bonding; Isomerism; Models, Molecular; Photochemistry; Retinaldehyde | 2005 |
In vivo treatment with CPT-11 leads to differentiation of neuroblastoma xenografts and topoisomerase I alterations.
Topoisomerase I inhibitors, such as CPT-11, are potent anticancer drugs against neuroblastoma (NB). Differentiating agents, such as retinoids, improve the survival of children with metastatic NB. To characterize the biological effects associated with exposure to CPT-11 in vivo, athymic mice bearing a human NB xenograft, named IGR-NB8 and characterized as an immature NB with poor prognostic markers, were treated with CPT-11. Prolonged stable disease was observed, resulting in an overall tumor growth delay of 115 days. During treatment, tumors differentiated into ganglioneuroblastomas (GGNB), which reverted into an immature phenotype when treatment was discontinued. In contrast, 13-cis retinoic acid failed to induce differentiation of IGR-NB8 in vivo. Tumor differentiation was associated with decreased N-myc expression, induction of p73 expression in the perinuclear area and cytoplasm, and a dramatic 35-fold decrease in topoisomerase I (topo I) catalytic activity. The full-length Mr 100,000 topo I protein was present in both pre and post-treatment immature NB xenografts. In contrast, differentiated GGNBs did not contain the Mr 100,000 protein but an intense Mr 48,000 topo I fragment. Furthermore, redistribution of the Mr 48,000 and 68,000 forms to the cytoplasm was observed in differentiated tumors. The same pattern of topo I expression and catalytic activity was observed in NBs and GGNBs obtained from pediatric patients. Our data suggest that prolonged in vivo exposure to CPT-11 induces differentiation of NB xenografts, which is associated with truncation of the topo I enzyme, relocation of the degraded forms to the cytoplasm, and decreased catalytic activity. Topics: Animals; Antineoplastic Agents, Phytogenic; Camptothecin; Cell Differentiation; Cell Division; Child; Child, Preschool; Diterpenes; DNA Topoisomerases, Type I; Enzyme Inhibitors; Female; Humans; Infant; Infant, Newborn; Irinotecan; Male; Mice; Mice, Nude; Neuroblastoma; Retinaldehyde; Topoisomerase I Inhibitors; Xenograft Model Antitumor Assays | 2004 |
Enzymatic characterization of recombinant mouse retinal dehydrogenase type 1.
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 | 2003 |
All-trans to 13-cis retinal isomerization in light-adapted bacteriorhodopsin at acidic pH.
The flash photolysis kinetic spectra of the intermediate M(412) of bacteriorhodopsin were monitored during the process of acid titration. In the light-adapted state, the maximum peak amplitude of M(412) absorbance of bacteriorhodopsin decreased (pK(a)=3.40+/-0.05) as the pH decreased from 7.3 to 1.9. In the dark-adapted state, the maximum peak amplitude of M(412) absorbance of bacteriorhodopsin increased as the pH decreased from 6.9 to 4.1, and then decreased (pK(a)=2.85+/-0.05) as the pH dropped to 2.1. These different trends in the change in the maximum peak amplitude suggested that not only the transition of purple membrane to blue membrane had taken place in both light and dark-adapted states, but also the fraction of all-trans-bR had changed during the acid titration. The pH-dependent absorption changes at 640 nm of bacteriorhodopsin in both light- and dark-adapted states were also observed. The pK(a)-values of the purple-to-blue transition were 3.80+/-0.05 in light-adapted state and 3.40+/-0.05 in dark-adapted state, respectively. According to Balashov's method, the fraction of all-trans-bR was assayed as the pH decreased. All these results indicated that the purple-to-blue transition of light-adapted bacteriorhodopsin was accompanied by an all-trans to 13-cis retinal isomerization at acidic pH. Topics: Bacteriorhodopsins; Diterpenes; Halobacterium salinarum; Hydrogen-Ion Concentration; Isomerism; Kinetics; Light; Photolysis; Retinaldehyde; Spectrophotometry | 2002 |
Light-induced hydrolysis and rebinding of nonisomerizable bacteriorhodopsin pigment.
Bacteriorhodopsin (bR) is characterized by a retinal-protein protonated Schiff base covalent bond, which is stable for light absorption. We have revealed a light-induced protonated Schiff base hydrolysis reaction in a 13-cis locked bR pigment (bR5.13; lambda(max) = 550 nm) in which isomerization around the critical C13==C14 double bond is prevented by a rigid ring structure. The photohydrolysis reaction takes place without isomerization around any of the double bonds along the polyene chain and is indicative of protein conformational alterations probably due to light-induced polarization of the retinal chromophore. Two photointermediates are formed during the hydrolysis reaction, H450 (lambda(max) = 450 nm) and H430 (lambda(max) = 430 nm), which are characterized by a 13-cis configuration as analyzed by high-performance liquid chromatography. Upon blue light irradiation after the hydrolysis reaction, these intermediates rebind to the apomembrane to reform bR5.13. Irradiation of the H450 intermediate forms the original pigment, whereas irradiation of H430 at neutral pH results in a red shifted species (P580), which thermally decays back to bR5.13. Electron paramagnetic resonance (EPR) spectroscopy indicates that the cytoplasmic side of bR5.13 resembles the conformation of the N photointermediate of native bR. Furthermore, using osmotically active solutes, we have observed that the hydrolysis rate is dependent on water activity on the cytoplasmic side. Finally, we suggest that the hydrolysis reaction proceeds via the reversed pathway of the binding process and allows trapping a new intermediate, which is not accumulated in the binding process. Topics: Apoproteins; Bacteriorhodopsins; Diterpenes; Electron Spin Resonance Spectroscopy; Hydrogen-Ion Concentration; Hydrolysis; Isomerism; Kinetics; Light; Models, Molecular; Molecular Structure; Protein Binding; Retinaldehyde; Schiff Bases; Spectrophotometry | 2002 |
Kinetic analysis of mouse retinal dehydrogenase type-2 (RALDH2) for retinal substrates.
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 | 2002 |
Isomerization of all-trans-retinol to cis-retinols in bovine retinal pigment epithelial cells: dependence on the specificity of retinoid-binding proteins.
In the retinal rod and cone photoreceptors, light photoactivates rhodopsin or cone visual pigments by converting 11-cis-retinal to all-trans-retinal, the process that ultimately results in phototransduction and visual sensation. The production of 11-cis-retinal in adjacent retinal pigment epithelial (RPE) cells is a fundamental process that allows regeneration of the vertebrate visual system. Here, we present evidence that all-trans-retinol is unstable in the presence of H(+) and rearranges to anhydroretinol through a carbocation intermediate, which can be trapped by alcohols to form retro-retinyl ethers. This ability of all-trans-retinol to form a carbocation could be relevant for isomerization. The calculated activation energy of isomerization of all-trans-retinyl carbocation to the 11-cis-isomer was only approximately 18 kcal/mol, as compared to approximately 36 kcal/mol for all-trans-retinol. This activation energy is similar to approximately 17 kcal/mol obtained experimentally for the isomerization reaction in RPE microsomes. Mass spectrometric (MS) analysis of isotopically labeled retinoids showed that isomerization proceeds via alkyl cleavage mechanism, but the product of isomerization depended on the specificity of the retinoid-binding protein(s) as evidenced by the production of 13-cis-retinol in the presence of cellular retinoid-binding protein (CRBP). To test the influence of an electron-withdrawing group on the polyene chain, which would inhibit carbocation formation, 11-fluoro-all-trans-retinol was used in the isomerization assay and was shown to be inactive. Together, these results strengthen the idea that the isomerization reaction is driven by mass action and may occur via carbocation intermediate. Topics: Animals; Cattle; cis-trans-Isomerases; Diterpenes; Humans; Hydrochloric Acid; Isomerism; Mass Spectrometry; Mathematical Computing; Microsomes; Photochemistry; Pigment Epithelium of Eye; Retinaldehyde; Retinoids; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Retinyl Esters; Sodium Hydroxide; Vitamin A | 2000 |
Isomerization of all-trans-9- and 13-desmethylretinol by retinal pigment epithelial cells.
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 | 1999 |
Lipocalin-type prostaglandin D synthase (beta-trace) is a newly recognized type of retinoid transporter.
Lipocalin-type prostaglandin D synthase is responsible for the biosynthesis of prostaglandin D2 in the central nervous system and the genital organs and is secreted into the cerebrospinal fluid and the seminal plasma as beta-trace. Here we analyzed retinoids binding of the enzyme by monitoring the fluorescence quenching of an intrinsic tryptophan residue, and appearance of circular dichroism around 330 nm, and a red shift of the UV absorption spectra of retinoids. We found that the enzyme binds all-trans- or 9-cis-retinoic acid and all-trans- or 13-cis-retinal, but not all-trans-retinol, with affinities (Kd of 70-80 nM) sufficient for function as a retinoid transporter. All-trans-retinoic acid inhibited the enzyme activity in a noncompetitive manner, suggesting that it binds to the same hydrophobic pocket as prostaglandin H2, the substrate for prostaglandin D synthase, but at a different site in this pocket. It is likely that this enzyme is a bifunctional protein that acts as both retinoid transporter and prostaglandin D2-producing enzyme. Topics: Alanine; Alitretinoin; Animals; Biological Transport, Active; Circular Dichroism; Cystine; Diterpenes; Intramolecular Oxidoreductases; Isomerases; Isomerism; Kinetics; Lipocalins; Models, Molecular; Protein Conformation; Rats; Recombinant Proteins; Retinaldehyde; Retinoids; Retinol-Binding Proteins; Retinol-Binding Proteins, Plasma; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet; Tretinoin | 1997 |
Reason for the lack of light-dark adaptation in pharaonis phoborhodopsin: reconstitution with 13-cis-retinal.
The reconstitution of pharaonis phoborhodopsin was performed by incubation of its opsin with 13-cis-retinal. Spectrum change was very slow, and two phases of the change were observed: the first and second phases are due to the transient formation of 13-cis pigment and spontaneous isomerization to all-trans-retinal, respectively. Slow binding supports an idea that the retinal binding pocket of ppR is highly restricted. Being bent in the configuration, 13-cis-retinal cannot be accommodated in the pocket due to the steric hindrance. This is a possible reason for the lack of light-dark adaptation. Topics: Archaeal Proteins; Bacteriorhodopsins; Carotenoids; Darkness; Diterpenes; Halobacteriales; Halorhodopsins; Light; Photochemistry; Retinaldehyde; Sensory Rhodopsins; Spectrophotometry | 1995 |
Growth factors and hormones which affect survival, growth, and differentiation of the MCF-7 stem cells and their descendants.
The human breast tumor cell line was separated by Percoll density gradient centrifugation into six different subpopulations, A to F, one of which (E) appears to contain the stem cells on the basis of several criteria (M. Resnicoff et al. 1987, Proc. Natl. Acad. Sci. USA 84, 7295. We now analyzed the response of the isolated subpopulations to insulin, thrombin, PGF2 alpha, estradiol, and 13-cis-retinal. We demonstrate that the first two growth factors stimulate [3H]thymidine incorporation in the more differentiated subpopulations (D and F), while PGF2 alpha has mitogenic activity in subpopulations C and D. In the absence of any added growth factor, estradiol has the extreme and transient capacity of allowing the stem cell to detach from the tissue culture dish and to grow in suspension as multicellular aggregates (MCF-7/SE cells). 13-cis-Retinal acts as a negative modulator of differentiation and protects the cells from the inhibitory and differentiation activity of Na-butyrate. Topics: Breast Neoplasms; Butyrates; Butyric Acid; Cell Differentiation; Cell Division; Cell Survival; Culture Media; Dinoprost; Diterpenes; DNA; Estradiol; Hormones; Humans; Insulin; Neoplastic Stem Cells; Retinaldehyde; Retinoids; Thrombin; Tumor Cells, Cultured | 1989 |
Changes in the protonation state of bacterio-opsin during reconstitution of bacteriorhodopsin.
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 | 1980 |