Assigning the resonance Raman spectral features of rhodopsin, isorhodopsin and bathorhodopsin in bovine photostationary state spectra.

Abstract—High resolution resonance Raman spectra of rhodopsin. isorhodopsin and photostationary state mixtures containing a high percentage of bathorhodopsin arc presented. New spectral features are detected which were not obsei-ved in lower resolution studies by other workers. All of the hands in the photostationary state spcctra arc assigned based on pure rhodopsin and isorhodopsin resonance Raman results and alterations in the photostationary state mixture. The spectral features in these spectra are invariant from 20 to 150K indicating that retinal and protein structural alteration, consistent with a model of excitation proposed by Lewis, occurs in steady-state spectra even at 20 K. In addition, the relative intensity of certain features in the photostationary state spectra are altered upon D₂O suspension. One explanation for these alterations is that the contributions of various intcrmediates to the photostationary state mixture are changed when membrane fragments are suspended in D₂O.     

Ordering in a nematic side-chain polymer. A proton and deuterium nuclear magnetic resonance study

 The aim of this work is to study in detail the orientational structure of liquid-crystalline polymers with different molecular weight. The advantage of our approach is the use of broad-band ¹H and ²H NMR spectroscopy for the analysis of the orientational order of the main-chain and mesogenic groups in conjunction with small-angle neutron scattering results. Investigations of a series of partially deuterated side-chain polymers with methoxybenzylbenzoate mesogenic groups have been carried out. The deuteration of the main chain enables separate access to order and orientation effects of the side chain and the main chain. The molecular orientation of the substances investigated can be frozen in its nematic phase below the glass-transition temperature. It is independent of the molecular weight, whereas the dynamics of the director reorientation (rotational viscosity) in the nematic phase strongly depends on the molecular weight. The main-chain deuterons provide powder spectra after orientation of the polymer in magnetic fields, indicating a very weak or lacking orientational anisotropy. A complete decoupling of the main and the side group can be deduced. Received: 15 September 1999 Accepted: 7 February 2000     

Absorption of schiff-base retinal chromophores in vacuo

The absorption spectrum of the all-trans retinal chromophore in the protonated Schiff-base form, that is, the biologically relevant form, has been measured in vacuo, and a maximum is found at 610 nm. The absorption of retinal proteins has hitherto been compared to that of protonated retinal in methanol, where the absorption maximum is at 440 nm. In contrast, the new gas-phase absorption data constitute a well-defined reference for spectral tuning in rhodopsins in an environment devoid of charges and dipoles. They replace the misleading comparison with absorption properties in solvents and lay the basis for reconsidering the molecular mechanisms of color tuning in the large family of retinal proteins. Indeed, our measurement directly shows that protein environments in rhodopsins are blue- rather than red shifting the absorption. The absorption of a retinal model chromophore with a neutral Schiff base is also studied. The data explain the significant blue shift that occurs when metharhodopsin I becomes deprotonated as well as the purple-to-blue transition of bacteriorhodopsin upon acidification.     

Synthesis of deuterium labeled ezetimibe and its glucuronide conjugate

Journal of Radioanalytical and Nuclear Chemistry - Ezetimibe is an effective cholesterol absorption inhibitor approved for the combined treatment with statins to reduce LDL-C level. Ezetimibe...     

Absorption studies of neutral retinal Schiff base chromophores

The neutral retinal Schiff base is connected to opsin in UV sensing pigments and in the blue-shifted meta-II signaling state of the rhodopsin photocycle. We have designed and synthesized two model systems for this neutral chromophore and have measured their gas-phase absorption spectra in the electrostatic storage ring ELISA with a photofragmentation technique. By comparison to the absorption spectrum of the protonated retinal Schiff base in vacuo, we found that the blue shift caused by deprotonation of the Schiff base is more than 200 nm. The absorption properties of the UV absorbing proteins are thus largely determined by the intrinsic properties of the chromophore. The effect of approaching a positive charge to the Schiff base was also studied, as well as the susceptibility of the protonated and unprotonated chromophores to experience spectral shifts in different solvents.     

Induced orientation of free chains in a strained rubber: A deuterium magnetic resonance study

Linear free polydimethylsiloxane (PDMS) chains are dissolved in a PDMS model network. When the system is uniaxially stretched, the free chains, as well as the network chains themselves, acquire a uniaxial orientational order, which is monitored by ²H NMR. The induced orders measured on both kinds of chains are comparable. The order is then investigated as functions of the free chain length and concentration.     

A resonance Raman study of the higher-lying electronic states of styrene vapor

Resonance Raman spectra of styrene vapor excited in the S₂ /S₃ and S₄ absorption systems are reported. The lack of double bond torsional scattering activity is taken as evidence of high barriers to rotation in all the excited states examined. Consequences for styrene photoisomerization are discussed. Excitation to S₂/S₃ is delocalized in nature, while S₄ excitation is benzene-like. The observed scattering intensities are largely in agreement with the jet absorption analysis and the results of Franck-Condon calculations.     

Preparation of l-serine and l-cystine stereospecifically labeled with deuterium at the β-position

The synthesis of l-serine and l-cystine stereospecifically labeled with deuterium at the β-position is described. The carboxyl group of d-serine was transformed into chirally deuterium-labeled alcohol via asymmetric reduction of 1-deuterio aldehyde, while the original hydroxymethyl group was converted into a carboxyl functionality to afford (2S,3R)-[3-²H]serine. Functional group interconversions of the hydroxyl group in the obtained deuterium-labeled l-serine gave (2R,2′R,3S,3′S)-[3,3′-²H₂]cystine.     

The mechanism of formation of volatile hydrides by tetrahydroborate(III) derivatization: A mass spectrometric study performed with deuterium labeled reagents

In order to clarify the mechanism of hydride formation, the isotopic composition of arsine, stibine, bismuthine, germane, stannane and hydrogen selenide formed by derivatization with either NaBD₄ (TDB) or NaBH₄ (THB) with inorganic As(III), Sb(III), Bi(III), Ge(IV), Sn(IV) and Se(IV) in aqueous reaction media and under various reaction conditions was determined. Batch hydride generation and gas chromatography–mass spectrometry (GC–MS) were employed. The analyte, present in 0.5–5 ml of acid solution (0.1–10 M in HCl or HNO₃ or HClO₄) was derivatized with 1 ml of 0.25–0.5 M TDB / THB in 0.1 M NaOH solution. For TDB derivatization in H₂O reaction media, almost pure BiD₃ and SbD₃ were always obtained for Bi(III) and Sb(III). Nearly pure AsD₃ could be obtained only under some reaction conditions. In general, for As(III), the isotopic composition of the arsines depends strongly on reaction conditions and included all possible AsH_nD_3−n from almost pure AsD₃ to almost pure AsH₃. For Ge(IV) and Sn(IV), the isotopic composition of generated GeH_nD_4−n and SnH_nD_4−n depends on reaction conditions, but pure GeD₄ and SnD₄ could never be obtained. Pure H₂Se was obtained in all cases, independent of reaction conditions. The occurrence of side reactions involving D–H exchange in TBD during its hydrolysis and before the derivatization step, as well as on recently formed hydrides following derivatization was investigated. D–H exchange in TDB during acid hydrolysis appears to occur to a limited extent. Amongst the hydrides, H₂Se undergoes H–D exchange whereas germane and stannane do not exchange at all. Arsine undergoes D–H exchange at elevated acidities (pH < 0) whereas stibine and bismuthine do not exchange significantly during the generation and stripping steps.A reaction model for hydride generation is proposed accounting for primary reactions giving rise to hydride formation through reaction intermediates, as well as side reactions involving D–H exchange and decomposition of reactive hydroboron species, reaction intermediates and final products. Hydrides are formed by direct hydrogen transfer from boron to the analyte atom, most likely through concerted mechanisms taking place via reaction intermediates.     

A deuterium nuclear quadrupole double resonance study of the ferroelectric phase transition in potassium dihydrogen phosphate

A study has been made of the nuclear quadrupole double resonance (NQDR) spectrum of deuterium in a powdered sample of potassium dihydrogen phosphate, KH₂PO₄ or KDP for short, both as a function of temperature and of percentage deuteration. Double transitions have also been observed and these enable the orientation of the ferroelectric electric field gradient (EFG) tensor to be determined.     

A resonance Raman,surface-enhanced resonance Raman,IR, and ab initio vibrational spectroscopic study of nickel(II) tetraazaannulene complexes

The IR and resonance Raman spectra of the nickel(II) complexes of dibenzo[b,i][1,4,8,11]tetraaza[14]annulene (TAA) and 5,7,12,14-tetramethyldibenzo[b,i][1,4,8,11]tetraaza[14]annulene (TMTAA) have been measured and compared with ab initio calculations of the vibrational wavenumbers at the B3-LYP level using the LanL2DZ basis set. An excellent fit is found between the experimental and calculated data, enabling precise vibrational assignments to be made. Surface-enhanced resonance Raman spectra were obtained following adsorption on Ag electrodes, with potentials in the range -0.1 to -1.1 V vs Ag/AgCl. There is evidence for contributions from both the electromagnetic and charge transfer (CT) surface enhancement mechanisms. The data indicate that variations in band intensities with electrode potential can be interpreted in terms of the CT mechanism.     

Structural anisotropy study by resonance Raman scattering in nematics with different dyes

Ionic liquid crystals based on congruent ion pairs composed of mesogenic cations and anions of similar shape provide an attractive tool for the tuning of mesophase properties. Here, the effect of the number and symmetry of lipophilic side chains and the type of head group on the phase type and thermal mesophase properties was probed by the synthesis and investigation of two series of novel guanidinium and imidazolium sulphonates and compared with the corresponding iodides. Their mesomorphic properties were examined via differential scanning calorimetry, optical polarising microscopy and X-ray diffraction. While derivatives bearing only one alkoxy chain in either cation or anion with up to three alkoxy chains in total within the ion pairs display smectic A mesophases, hexagonal columnar mesophases were observed for all other compounds with four or five alkoxy chains totally irrespective of the head group. However, with increasing steric bulk, i.e. with a total of six alkoxy chains, the symmetry of the aryl moiety in the anion as well as the type of head group becomes relevant, resulting in hexagonal columnar or plastic phases for guanidinium sulphonates with symmetrical anions, while those with unsymmetrical anions were non-mesomorphic. In contrast, the corresponding imidazolium sulphonates displayed cubic phases for the combinations of symmetrical cation/symmetrical anion and symmetrical cation/unsymmetrical anion. If both ions are unsymmetrically substituted, the imidazolium sulphonate displayed a hexagonal columnar phase. The results further demonstrate the utility of the congruent ion pairs for tailor-made ionic liquid crystals.     

Double-quantum 13C nuclear magnetic resonance of bathorhodopsin, the first photointermediate in mammalian vision

The 13C chemical shifts of the primary visual photointermediate bathorhodopsin have been observed by performing double-quantum magic-angle-spinning NMR at low temperature in the presence of illumination. Strong isomerization shifts have been observed upon the conversion of rhodopsin into bathorhodopsin.     

Structural anisotropy study by resonance Raman scattering in nematics with different dyes

The method for the structural anisotropy determination, which is based on a comparison of the degree of polarization of absorption and resonance Raman bands, is tested with three dyes having different molecular dimensions in one nematic liquid crystal. The experimentally observed correlation between the variation of the structural anisotropy factor and the order parameters deviates from the predictions of the Segre model. This method could also be applied to determine the local field anisotropy for liquid crystals, but we must take into account the observed dependence of the structural anisotropy on the dimensions of the probe molecules.     

Synthesis of deuterium labeled compounds by KCN-assisted hydrolysis of phosphonium salts

We developed a facile deuterium labeling method for benzylic or allylic halides or acetates systems. Conversion of the halides or acetates to the corresponding phosphonium salts and the following mild hydrolysis with KCN afforded the deuterium labeled compounds in good yields.     

Interaction of hypericin with serum albumins: surface-enhanced Raman spectroscopy, resonance Raman spectroscopy and molecular modeling study

Surface-enhanced Raman spectroscopy, resonance Raman spectroscopy and molecular modeling were employed to study the interaction of hypericin (Hyp) with human (HSA), rat (RSA) and bovine (BSA) serum albumins. The identification of the binding site of Hyp in serum albumins as well as the structural model for Hyp/HSA complex are presented. The interactions mainly reflect: (1) a change of the strength of H bonding at the N1-H site of Trp; (2) a change of the Trp side-chain conformation; (3) a change of the hydrophobicity of the Trp environment; and (4) a formation of an H-bond between the carbonyl group of Hyp and a proton donor in HSA and RSA which leads to a protonated-like carbonyl in Hyp. Our results indicate that Hyp is rigidly bound in IIA subdomain of HSA close to Trp214 (distance 5.12 A between the centers of masses). In the model presented the carbonyl group of Hyp is hydrogen bonded to Asn458. Two other candidates for hydrogen bonds have been identified between the bay-region hydroxyl group of Hyp and the carbonyl group of the Trp214 peptidic link and between the peri-region hydroxyl group of Hyp and the Asn458 carbonyl group. It is shown that the structures of the Hyp/HSA and Hyp/RSA complexes are similar to, and in some aspects different from, those found for the Hyp/BSA complex. The role of aminoacid sequence in the IIA subdomains of HSA, RSA and BSA is discussed to explain the observed differences.     

The synthesis of deuterium labeled macrocyclic polyethers (crown ethers)

Five tetradeuterated analogues ( 1a, 1b, 1c, 1d , and 1e ) of 3,6,9,12,15-pentaoxa-21-aza-bicyclo[15.3.1]heneicosa-1(21),17,19-triene (pyridyl-18-crown-6, 1 ) have been prepared corresponding to the five unique protonated carbon atoms of the polyether cycle. Deuterium labeling was ≥ 99% for four of the compounds and ≥ 97% for the other (1a ).