Transitions Of Chromophores Research Articles

Structure analyses of biomolecules using a synchrotron radiation circular dichroism spectrophotometer

The vacuum-ultraviolet circular dichroism (VUVCD) spectra of various saccharides and proteins were measured in aqueous solution down to 160 nm, using a VUVCD spectrophotometer at the Hiroshima Synchrotron Radiation Center (HSRC). Most mono- and disaccharides exhibit characteristic peaks at around 170 nm that are sensitive to the anomeric and axial/equatorial configurations of hydroxyl groups, trans– gauche conformations of hydroxymethyl group, and the type of glycosidic linkage. The VUVCD spectra of proteins below 190 nm exhibit characteristic bands corresponding to the different secondary structures. Predicting the content and number of α-helix and β-strand segments is greatly improved, when lower wavelength CD spectra are available. These results demonstrate that synchrotron radiation VUVCD spectroscopy is useful for structure analyses of biomolecules based on the higher energy transitions of chromophores including hydroxyl and acetal groups.

Synthesis and Characterization of Segmented Liquid Crystalline Polymers with the Azo Group in the Main Chain

The synthesis of 4'-hydroxyphenyl(4-hydroxy-3-allyl)azobenzene is reported. The molecule is mesogenic and it can be used for preparing nematic main-chain segmented polymers. The nematic phase is the result of the effect of the side allyl group which, while it destabilizes the crystalline phase, does not affect the occurring of the nematic order. In the crystalline state, a large hexagonal unit cell containing five to six chains is observed for polymers with longer flexible spacer in the chain. Chloroform solutions and thin films of polymers absorb in the UV-visible range, at approximately 332 and 439 nm, corresponding to the ﷿ f ﷿* and n f ﷿* electronic transitions of azoaromatic chromophores, respectively. Under appropriate UV irradiation we have obtained the photoisomerizations trans-cis and cis-trans, which are fully reversible in the case of solutions. In the solid state a photostationary state is reached.

A general and nonempirical approach to the determination of the absolute configuration of 1-aryl-1,2-diols.

We describe herein a simple, general, and reliable nonempirical approach, based on the exciton coupling method, to assign the absolute configuration of the benzylic stereogenic center of 1-aryl-1,2-diols. According to this method, it is only necessary to prepare the 4-biphenylboronic esters of the diols and to record their CD spectra in the 230-300 nm range, i.e., in the range corresponding to the long-axis (1)L(a) transition of the biphenyl chromophore. From the sign of the CD couplet or Cotton effect at 260 nm it is possible to know the chirality defined by the aryl and biphenyl chromophore transitions and then to determine the absolute configuration of the benzylic carbon. By this approach, simple rules have been formulated which allow us to establish the absolute configuration of many classes of 1-aryl-1,2-diols.

Electronic spectra and structures of d2 molybdenum-oxo complexes. Effects of structural distortions on orbital energies, two-electron terms, and the mixing of singlet and triplet states.

Molybdenum-oxo ions of the type [Mo(IV)OL(4)Cl](+) (L = CNBu(t), PMe(3), (1)/(2)Me(2)PCH(2)CH(2)PMe(2)) have been studied by X-ray crystallography, vibrational spectroscopy, and polarized single-crystal electronic absorption spectroscopy (300 and ca. 20 K) in order to investigate the effects of the ancillary ligand geometry on the properties of the MotriplebondO bond. The idealized point symmetries of the [Mo(IV)OL(4)Cl](+) ions were established by X-ray crystallographic studies of the salts [MoO(CNBu(t)())(4)Cl][BPh(4)] (C(4)(v)), [MoO(dmpe)(2)Cl]Cl.5H(2)O (C(2)(v)), and [MoO(PMe(3))(4)Cl][PF(6)] (C(2)(v)()); the lower symmetries of the phosphine derivatives are the result of the steric properties of the phosphine ligands. The Motbd1;O stretching frequencies of these ions (948-959 cm(-)(1)) are essentially insensitive to the nature and geometry of the equatorial ligands. In contrast, the electronic absorption bands arising from the nominal d(xy)() --> d(xz), d(yz) (n --> pi(MoO)) ligand-field transition exhibit a large dependence on the geometry of the equatorial ligands. Specifically, the electronic spectrum of [MoO(CNBu(t)())(4)Cl](+) exhibits a single (1)[n --> pi(xz)(,)(yz)] band, whereas the spectra of both [MoO(dmpe)(2)Cl](+) and [MoO(PMe(3))(4)Cl](+) reveal separate (1)[n --> pi(xz)] and (1)[n --> pi(yz)] bands. A general theoretical model of the n --> pi state energies of structurally distorted d(2) M(triplebondE)L(4)X chromophores is developed in order to interpret the electronic spectra of the phosphine derivatives. Analysis of the n --> pi transition energies using this model indicates that the d(xz) and d(yz) pi(MotriplebondO) orbitals are nondegenerate for the C(2)(v)-symmetry ions and the n --> pi(xz) and n --> pi(yz) excited states are characterized by different two-electron terms. These effects lead to a significant redistribution of intensity between certain spin-allowed and spin-forbidden absorption bands. The applicability of this model to the excited states produced by delta --> pi and pi --> delta symmetry electronic transitions of other chromophores is discussed.

Non-photochemical versus photochemical hole burning in hyperquenched glassy water and cubic ice

The Shu–Small mechanism (J. Opt. Soc. Am. B 9 (1992) 724) for non-photochemical hole burning (NPHB) of electronic transitions of chromophores in amorphous hosts at low temperatures is based on a hierarchy of configurational tunneling events that begin in the outer shell and terminate in the inner shell that surrounds the chromophore. Connectivity between the intrinsic and extrinsic two level systems that stem from structural disorder and the excess free volume of glasses are key features of this mechanism. This mechanism has recently been modified to take into account multilevel extrinsic systems (J. Chem. Phys. 114 (2001) 9105). An important prediction of the Shu–Small mechanism is that NPHB should cease upon formation of a crystalline phase produced, for example, by warming of a vitrified glass. In this paper data are presented that are consistent with that prediction. The systems studied are aluminum phthalocyanine tetrasulphonate (APT) and free base phthalocyanine tetrasulphonate (PcT) in hyperquenched glassy films of water (HGW) and cubic ice ( I c). The hole-burning mechanisms for APT and PcT are non-photochemical and photochemical, respectively. Surprisingly, the hole-growth kinetics for PcT are as highly dispersive as are the kinetics for APT. This means that the kinetics for photoinduced proton tautomerization in PcT are strongly influenced by its nanoenvironment of water molecules, some of which are involved in H-bonding to the –NH groups of PcT.

Changes in the retinal transition dipole moment in bacteriorhodopsin of the purple membrane of Halobacterium Salinarium at the so-called PH(rev).

The chromophore transition dipole moment of light-adapted wild-type bacteriorhodopsin (WT-bR) in suspension is evaluated for pH range of 2.8-12. Significant variations in the transition dipole moment are observed at the so-called pH(rev). The results have reported a value of 11 and 8.5 Debye for chromophore transition dipole moment of the neutral purple and acid blue form of bacteriorhodopsin, respectively. There may be a correlation with the process of reversing the direction of the permanent electric dipole moment, due to the reversal of the surface charge asymmetry, of the purple membrane to its opposite side at that pH(rev).

Simple and accurate computations of solvatochromic shifts in pi --> pi* transitions of aromatic chromophores.

A new approach is introduced for calculating the spectral shifts of the most bathochromic pi --> pi* transition of an aromatic chromophore in apolar environments. As an example, perylene in solid and liquid n-alkane matrixes was chosen, and all shifts were calculated relative to one well-defined solid-inclusion system. It was shown that a simple two-level treatment of the solute using Hückel theory yields spectral shifts in excellent agreement with experimental results for the most prominent inclusion sites of perylene in solid n-alkane surroundings and for the dilute solutions in liquid n-alkanes. The idea is general enough to be applied to any aromatic chromophore in a nonpolar solvent matrix. In contrast to earlier treatments, this approach is based on geometry- and environment-dependent polarizabilities, employs an r(-4) dependence for the dispersion energy, and is conceptually very simple and computationally very efficient.

Ligand Substituents of Ruthenium Dipyridophenazine Complexes Sensitively Determine Orientation in Liposome Membrane

A series of ruthenium(II) complexes comprising differently substituted dipyridophenazine (dppz) ligands have been examined with respect to their orientation when bound to phosphatidylcholine liposome bilayers. The liposomes were subjected to shear flow, thereby deformed into ellipsoid shapes, and the orientation of chromophores in the bilayers could then be examined with polarized light (linear dichroism, LD). After correction for turbidity dichroism, LD distinguishes between chromophore transition moments oriented along the lipid chains (negative LD), and parallel to the membrane surface (positive LD). The polarizations, energies, and absorption overlap of the electronic transitions of the ruthenium complexes, known from previous work, could be used to characterize the orientation of liposome bound complexes. From luminescence lifetime studies further information about the localization of the chromophores was obtained. Assuming that the positively charged metal moiety of the complexes remains close to th...

Understanding the Optical Band Shape: Coumarin-153 Steady-State Spectroscopy

We have developed a band-shape analysis of optical transitions in polarizable chromophores characterized by large magnitudes of the transition dipole (intense transitions). The model is tested on steady-state spectra of the coumarin-153 optical dye, employing an explicit solvent description accounting for dipole moment, quadrupole moment, and polarizability of the solvent molecules. The calculations are performed for solvents ranging from nondipolar to strongly dipolar. The solvent dependence of both the experimental Stokes shift and the spectral width is satisfactorily reproduced over the whole polarity range. The optical width is shown to demonstrate a qualitatively different solvent dependence for absorption and emission. The solvent-induced absorption width increases with solvent polarity, whereas the solvent-induced emission width passes through a maximum. This is a result of non-Gaussian statistics of the energy gap fluctuations in polarizable/electronically delocalized chromophores. The total (i.e., solvent and vibrational) emission width tends to pass through a broad maximum at low solvent polarities, decreasing with solvent polarity for highly polar solvents. This results from the combined influence of a solvent-induced mixing of the vacuum adiabatic states and a decrease of the vibrational reorganization energy with increasing solvent polarity. The latter effect arises as a result of a coupling of the vibrational and solvent nuclear modes due to the electronic state occupation number difference, making the vibrational reorganization energy solvent-dependent. The study revels a breakdown of the linear relation between the solvent-induced width and Stokes shift. The model suggests that the Franck−Condon factor of intense optical lines should significantly depend on the magnitude of the transition dipole.

Multicolored Protein Conformation States in the Photocycle of Transducer-Free Sensory Rhodopsin-I

Sensory rhodopsin-I (SRI), a phototaxis receptor of archaebacteria, is a retinal-binding protein that exists in the cell membrane intimately associated with a signal-transducing protein (HtrI) homologous to eubacterial chemotaxis receptors. Transducer-free sensory rhodopsin-I (fSRI), from cells devoid of HtrI, undergoes a photochemical cycle kinetically different from that of native SRI. We report here on the measurement and analysis of the photochemical kinetics of fSRI reactions in the 350–750-nm spectral range and in a 10 −7 s to 1 s time window. The lack of specific intermolecular interactions between SRI and HtrI results in early return of the ground form via distinct branching reactions in fSRI, not evident in the photocycle of native SRI. The chromophore transitions are loosely coupled to protein structural transitions. The coexistence of multiple spectral forms within kinetic intermediates is interpreted within the concept of multicolored protein conformational states.

Absorption spectra of chlorophyll a and b in Lhcb protein environment.

The spectral forms of the two chlorophyll species in higher plant Photosystem II antenna proteins have been experimentally determined within their protein environment. Recombinant CP29 and LHC II antenna proteins missing individual chromophores were obtained by over-expression in bacteria without any changing of the primary protein sequence and in vitro reconstitution. Difference absorption spectroscopy with respect to the corresponding proteins binding the complete pigment complement yielded the spectral shape and extinction of single chlorophyll a and b. A functional relation of their absorption was given by Gaussian subband decomposition covering the entire Q(x) and Q(y) optical region together with the absolute value of the molar extinction coefficient. With respect to analogous determinations reported in the literature for organic solvents, this information is valuable for further understanding the in-protein chlorophyll excited states and excited state dynamics: in particular, for the calculation of Förster transfer rates by means of chlorophyll-chlorophyll overlap integral employing the Stepanov relation for emission and single chromophore transition energies according to the results of mutational analysis of chlorophyll binding sites [Bassi et al. (1999) Proc Natl Acad Sci USA 96: 10056-10061; Remelli et al. (1999) J Biol Chem 274: 33510-33521].

Optical response functions for condensed systems with linear and quadratic electron–vibration coupling

Understanding the similarities and differences between optical coherence loss of electronic transitions of chromophores in glasses and in the glass forming solvent requires, in part, linear response (2-point correlation) functions, J(t;T). An approximate excited state vibrational Hamiltonian (He) which accounts for both linear and quadratic electron–phonon coupling is derived that is acceptable for mode frequency changes smaller than 30%. The associated linear response function for the case of no damping is obtained. A response function that includes damping is proposed for systems whose modes are either linearly or quadratically coupled. It is the product of three response functions, two of which are phononic and associated with linear and quadratic modes. The third response function is electronic with a dephasing frequency γel that is the width of the zero-phonon line. The total response function yields single-site absorption spectra in which folding of the widths of multi-phonon and sequence transitions occurs. Applications of the new response functions are made to the temperature dependence of single-site absorption and hole-burned spectra of the special pair band of the bacterial reaction center and the temperature dependence of the single site absorption spectrum of Al-phthalocyanine tetrasulphonate in glassy ethanol.

Volume and Enthalpy Changes upon Photoexcitation of Bovine Rhodopsin Derived from Optoacoustic Studies by Using an Equilibrium between Bathorhodopsin and Blue‐Shifted Intermediate

AbstractA complex multistep model including an equilibrium was used for the first time to analyze data obtained by laser‐induced optoacoustic spectroscopy (LIOAS) with bovine rhodopsin in washed membranes (T = 5–32°C). The thermal equilibrium was between bathorhodopsin, batho, and the blue‐shifted intermediate (BSI), i.e., batho ⇄ BSI. This analysis considers a 14‐ns intermediate detected by LIOAS at 25°C (with similar decay activation parameters as those reported for BSI in solubilized rhodopsin). The energy content of lumirhodopsin (lumi) and the structural volume difference between lumi and rhodopsin were essentially the same (Elumi. = 80 kJ/mol and ΔVlumi–rho = 40 mL/mol) for both a model based on a simple batho → BSI → lumi sequence and one including the batho ⇄; BSI equilibrium. However, the structural volume changes for the individual steps batho → BSI and BSI → lumi were markedly changed by the introduction of the equilibrium. With the rate constant for the forward reaction batho → BSI for solubilized rhodopsin determined by Hug et al. (Hug, S.J.; Lewis, W.J.; Einterz, C.M.; Thorgeirsson, T.E.; Kliger, D.S. Biochemistry, 1990, 29, 1475–1485) the structural volume changes obtained with the equilibrium model are ΔVBSI‐batho = 79 mL/mol and ΔVlumi‐BSI = −44 mL/mol. The resulting batho → BSI expansion is larger than with the sequential model, and ΔVlumi‐BSI is now a contraction instead of an expansion. A small expansion of 5 mL/mol is derived for the formation of batho with both models. The estimated energy content of BSI is clearly higher than the value of lumi. The structural volume changes between batho, BSI, and lumi derived from the equilibrium model correlate better than those from the simple sequential model with the chromophore transition dipole moment changes determined in the literature using linear dichroism measurements. The structural volume changes are discussed in terms of the variations upon transformation of the chromophore–protein interactions.

Quantitative calculations of fluorescence polarization and absorption anisotropy kinetics of double- and triple-chromophore complexes with energy transfer

A new method is presented for calculation of the fluorescence depolarization and kinetics of absorption anisotropy for molecular complexes with a limited number of chromophores. The method considers absorption and emission of light by both chromophores, and also energy transfer between them, with regard to their mutual orientations. The chromophores in each individual complex are rigidly positioned. The complexes are randomly distributed and oriented in space, and there is no energy transfer between them. The new "practical" formula for absorption anisotropy and fluorescence depolarization kinetics, P(t) = [3B(t) - 1 + 2A(t)]/[3 + B(t) + 4A(t)], is derived both for double- and triple-chromophore complexes with delta-pulse excitation. The parameter B(t) is given by (a) B(t) = cos2(theta) for double-chromophore complexes, and (b) B(t) = q12(t)cos2(theta 12) + q13(t)-cos2(theta 13) + q23(t)cos2(theta 23) for triple-chromophore complexes, where q12(t) + q13(t) + q23(t) = 1. Here theta ij are the angles between the chromophore transition dipole moments in the individual molecular complex. The parameters qij(t) and A(t) are dependent on chromophore spectroscopic features and on the rates of energy transfer.

STEADY‐STATE OPTICAL POLARIZATION ANISOTROPY OF RODLIKE MOLECULES UNDERGOING TORSIONAL TWISTING MOTIONS

Abstract An analytical expression was derived for the steady‐state optical polarization anisotropy expected for a rodlike molecule undergoing torsional twisting motions about its long molecular axis. This expression is parameterized in terms of the ratio τ/φ where τ is the excited‐state lifetime of the chromophore and Φ is a characteristic time for rotational twisting (Yoshimura et al., J. Mol. Biol. 179, 453–467, 1984) that is proportional to the torsional rigidity of the flexible rodlike molecule; the amplitude at any given value of τ/φ is also determined by the angle between the chromophore transition dipole and the long molecular axis of the molecule.

A cellular ligand-field model forl-lspectral intensities

The model for d-d or f-f intensities of ligand-field transitions in acentric chromophores was presented in part I. Here, that model is extended to include centrosymmetric systems in which intensity is deemed to arise from parity mixing induced during ungerade molecular vibrations. The same cellular structure is maintained for the parametrization scheme, although time-averaged t variables for each vibrational mode subsume local electronic properties and ligand displacements within each vibrational normal mode. The L t λ parameters of the ‘static’ model are replaced in the dynamic regime by L t α λ where α = x or y for local bending displacements normal to the metal-ligand (z) vector and α = z for pure stretching displacements. The physical and bonding significance of static- and dynamic-type parameters are described and compared.

Vibronic spectroscopy of jet-cooled hydrogen-bonded clusters

Mass-selected, two-photon resonant ionisation spectra of supersonically cooled p-cresol · (H 2O) n and phenol · (H 2O) n are reported. Cluster spectra with one, two and three water molecules attached can be unambiguously assigned. A monotonic shift of the electronic spectra with increasing cluster size is not observed here. The spectrum of p-cresol · (H 2O) 1, is red-shifted relative to the free p-cresol spectrum, while the p-cresol · (H 2O) 2,3 electronic origins are blue-shifted relative to p-cresol · (H 2O) 1 but still lie on the red side of the monomer. Simple, highest occupied molecular orbital—lowest unoccupied molecular orbital (HOMO—LUMO) considerations based on an initio calculations show that this can be explained by the inductive effect exerted on the O-atom of p-cresol, which acts as proton donor and acceptor in the H-bonding. The blue shift of n-π* transitions of H-bonded chromophores with carbonyl groups like CH 2O · (H 2O) 1 can be explained similarly. Vibrational spectra of supersonically cooled complexes of p-cresol with H 2O and CH 3OH were further analysed by dispersed fluorescence and stimulated emission, detected by two-colour ionisation dip. In p-cresol · (H 2O) 1, progressions of the intermolecular cluster stretch vibration and its combination bands with intramolecular cluster vibrations were observed with similar frequencies in the S 0 and S 1 states. In p-cresol · (H 2O) 3, further intense intermolecular bands arise, namely the hydrogen-bridge bending and torsion vibrations. This can be attributed to the lower symmetry of these clusters. Ab initio quantum chemical calculations show p-cresol · (H 2O) 3 to have a higher H-bond stretch frequency than p-cresol · (H 2O) 1, because its (unsymmetric) cyclic structure is more rigid. A characteristic pattern of CH 3 torsional bands in p-cresol is observed with complex structures due the CH 3 torsion—overall rotation interaction. Although their appearance is similar, the spacing of the monomer and cluster lines is very different, giving direct information on cluster structures from the rotational constants and moment of inertia.

DYNAMIC LINEAR DICHROISM IN CHROMOPROTEINS

Abstract—The time‐dependent polarized fluorescence and optical density transients arising from electronic excitation transport are derived for solutions of randomly oriented chromoproteins in which the chromophore transition moments have well‐defined orientations with respect to the protein. The treatment is extended to oriented monolayers of chromoproteins, in which the molecules are aligned with a particular protein‐fixed axis perpendicular to the surface plane and with random azimuthal angles about the surface normal. The initial ratio of parallel to perpendicular transients is invariably 3:1, irrespective of system dimensionality and chromophore organization. The residual anisotropy observed at long times is directly related to the relative orientations of the donor and acceptor chromophores. These calculations form a basis for detailed interpretation of ultrafast fluoresence and pump‐probe linear dichroism studies in photosynthetic systems, for which 3‐dimensional structures are becoming increasingly available.

Orientation of the protonated retinal Schiff base group in bacteriorhodopsin from absorption linear dichroism

Linear dichroism experiments are performed on light-adapted bacteriorhodopsin (BR568) films containing native retinal (A1) and its 3,4-dehydroretinal (A2) analogue to measure the angle between the chromophore transition dipole moment and the membrane normal. QCFF/pi calculations show that the angle between the transition moment and the long axis of the polyene is changed by 3.4 degrees when the C3-C4 bond is unsaturated. The difference vector between the two transition moments points in the same direction as the Schiff base (N----H) bond for the all-trans BR568 chromophore. Because the plane of the chromophore is perpendicular to the membrane plane, a comparison of the transition moment orientations in the A1- and A2-pigments enables us to determine the orientation of the N----H bond with respect to the absolute chromophore (N----C5 vector) orientation. The angles of the transition moments are 70.3 degrees +/- 0.4 degrees and 67.8 degrees +/- 0.4 degrees for the A1- and A2-pigments, respectively. The fact that the change in the transition moment angle (2.5 degrees) is close to the predicted 3.4 degrees supports the idea that the chromophore plane is nearly perpendicular to the membrane plane. The decreased transition moment angle in the A2-analogue requires that the N----H bond and the N----C5 vector point toward the same membrane surface. Available results indicate that the N----C5 vector points toward the exterior in BR568. With this assignment, we conclude that the N----H bond points toward the exterior surface and its most likely counterion Asp-212. This information makes possible the construction of a computer graphics model for the active site in BR568.

Orientation of the chromophore transition moment in the 4-leaved shape model for pea phytochrome molecule in the red-light absorbing form and its rotation induced by the phototransformation to the far-red-light absorbing form

Orientation of the chromophore transition moment in pea phytochrome (subunit molecular mass, 114 kDa) in the red-light-absorbing form (P r) and its rotation induced by the phototransformation to the far-red-light-absorbing (P fr) form was studied by measuring linear dichroism of oriented phytochrome prepared by a ‘gel squeezing method’. The phytochrome in P r showed a negative linear dichroism. On the contrary, the P fr form showed positive linear dichroism. Assuming that the molecular axis for the two-fold rotational symmetry in the ‘4-leaved shape model’ proposed recently for the dimeric molecular structure of the phytochrome [(1989) FEBS Lett. 247, 139–142], is oriented in parallel with the direction of the external constraint, the average azimuthal angle between the molecular axis and the transition moment of the chromophore was calculated as 59° or 121° for P r and 52° or 128° for P fr. These results explain well the polarotropic responses by phytochrome observed in plant organisms.