Studies of the Energy Transfer among Allophycocyanin from Phycobilisomes of Polysiphonia urceolata by Time-Resolved Fluorescence Isotropic and Anisotropic Spectroscopy

Abstract— The excitation energy transfer processes in the allophycocyanin (APC) monomer and trimer from phycobilisomes of Polysiphonia urceolata were studied using picosecond time-resolved fluorescence isotropic and anisotropic spectroscopy. Based on our experimental results, conclusions could be drawn as follows: (1) After the processes of exciton localization are finished, the localized excitation energy on any chromophore can be transferred to the other chromophores due to the weak couplings between them, and the processes among three p₈₄-phycocyaninbilin (PCB) chromophores in the center of the ring shape of the APC trimer are more important than those of between a₈₄- and p₈₄-PCB chromophores in the same monomer. (2) The decay time constants of 95 ± 5 ps and 40 ± 5 ps components, observed by us in this work, were assigned to the excitation energy transfer or redistribution between α₈₄- and β₈₄-PCB chromophores in the same monomer of the APC trimer and among three β₈₄-PCB chromophores in the center of the ring shape of the APC trimer, respectively. Specifically, the assignment of the decay constants for the 40 ± 5 ps component was different from those of previous results. (3) Based on the model of Debreczeny, and using the fluorescence residual anisotropy r(∞) with a probing wavelength of 650 nm, the angles between the C₃ symmetry axis and transition dipoles of α₈₄- and -PCB chromophores were found to be φ_a84= 67° and φ_β84= 148°, respectively, which are in agreement with the prediction of the X-ray crystal structure of APC. (4) The results show that anisotropy decays, observed with the APC trimer, did exhibit a strongly probing wavelength dependence that did not show up in the monomer.     

FLUORESCENCE RELAXATION KINETICS AND QUANTUM YIELD FROM THE ISOLATED PHYCOBILIPROTEINS OF THE BLUE-GREEN ALGA NOSTOC SP. MEASURED AS A FUNCTION OF SINGLE PICOSECOND PULSE INTENSITY, I

Abstract— Phycobilisomes from the blue-green alga Nostoc sp. are known to contain the phycobiliproteins: c-phycoerythrin (c-PE), c-phycocyanin (c-PC) and four forms of allophycocyanin (APC I, II, III, and B). We have made a detailed study of the effects of the intensity of a single 6 ps excitation pulse on the decay kinetics and the yield of fluorescence in the individual isolated phycobiliproteins at pH 7 and 23°C. The risetime of the fluorescence of c-PE, c-PC and APC was > 12 ps. We found that the decay of the fluorescence was exponential at intensities of 10¹⁴ photons/cm² in all the phycobiliproteins; the lifetimes being 1552 ± 31ps for c-PE, 2111 ± 83ps for c-PC, 1932 ± 165ps for APC I, 1870 ± 90ps for APC II, 1816 ± 88ps for APC III, (1869 ± 62ps for the averaged APC's I, II, and III), and 2667 ± 233 ps for APC B. We also found that the fluorescence decay became non-exponential in c-PE at excitation intensities < 10¹⁴ photons/cm², but was exponential for all the other phycobiliproteins even at a pulse intensity of 10¹⁵ photons/cm². The relaxation times of c-PE and c-PC decreased with excitation intensity above 10¹⁴ photons/cm². For c-PE and c-PC the relative fluorescence vs excitation intensity was readily described by a relationship derived for a model in which exciton–exciton annihilation occurs. In APC the fluorescence yield and relaxation time were only slightly dependent on the excitation intensity. The results are interpreted to indicate the occurrence of singlet–singlet annihilation intramolecularly among the several phycobilin chromophores within the individual phycobiliprotein molecules in solution. The s to f transfer time is less than 12ps in c-PC.     

Optical properties of a polydiacetylene crystal: poly-[1,6-di(N-carbazolyl)-2,4-hexadiyne]

Normal incidence reflection spectra and Kramers-Kronig transforms are reported for a polydiacetylene crystal, DCHD (substituent group: N-carbazolymethyl). The lowest energy optical transition for the polymer backbone is found at 15 300 cm^?1, the lowest value obtained thus far for a polydiacetylene. The polymer chain and the carbazolyl substituent group may be treated as isolated chromophores - the former dominating the visible portion of the spectra and the latter dominating the UV portion. Polarization studies for the (001) and (10$\text{1}\text{?}$) faces of DCHD permit unambiguous assignment of the two lowest energy transitions in the carbazolyl group at 28 750 cm^?1 and 33 250 cm^?1 as short- and long-axis polarized, respectively.     

THE EFFECT OF PERIPHERAL SUBSTITUTION ON THE BATHOCHROMIC SHIFT OF THE Qy TRANSITION OF BACTERIOCHLOROPHYLL DIMERS; in vitro MODELS OF THE PROTEIN EFFECT ON THE SPECTRUM OF PIGMENT CENTERS IN THE LIGHT-HARVESTING COMPLEXES*

Abstract— The effect of chemical modifications in the side groups of the isocyclic ring V on the formation, optical absorption and circular dichroism of bacteriochlorophyll (Bchl) dimers was examined in a mixture of formamide and water containing TritonX–100 and variable amounts of pyridine. Substitution of the carbomethoxy group in the C13² position with a hydrogen atom, had no effect on the dimerization constant but increased the shift of the Q_y transition by 1000 cm^-1 with respect to the native Bchl. Substitution of the C13 hydrogen atom with OH decreased the shift of the Q_y transition by 400 cm^-1. The similarity between the spectra of the modified Bchl dimers and Bchl dimers in vivo indicates that protein binding to the side groups at Bchl dimers may profoundly affect the energy of their Q_y transition but have minor effects on the Q_y transitions of the monomelic Bchl.     

Solid‐Phase Synthesis of Peptide Libraries Combining α‐Amino Acids with Inorganic and Organic Chromophores

The synthesis of two series of peptidic chains composed of bis(terpyridine)ruthenium(II) acceptor units and organic chromophores (coumarin, naphthalene, anthracene, fluorene) by stepwise solid‐phase peptide synthesis (SPPS) techniques is described. The first series of dyads comprises directly amide linked chromophores, while the second one possesses a glycine spacer between the two chromophores. All dyads were studied by UV/Vis and NMR spectroscopy, steady‐state luminescence, luminescence decay and electrochemistry, as well as by DFT calculations. The results of these studies indicate weak electronic coupling of the chromophores in the ground state. Absorpion spectra of all dyads are dominated by metal‐to‐ligand charge‐transfer (MLCT) bands around 500 nm. The bichromophoric systems, especially with coumarin as organic chromophore, display additional strong absorptions in the visible spectral region. All complexes are luminescent at room temperature (³MLCT). Efficient quenching of the fluorescence of the organic chromophore by the attached ruthenium complex is observed in all dyads. Excitation spectra indicate energy transfer from the organic dye to the ruthenium chromophore.     

A fluorescent turn-on detection scheme for α-fetoprotein using quantum dots placed in a boronate-modified molecularly imprinted polymer with high affinity for glycoproteins

This article describes a fluorescent molecularly imprinted polymer (MIP) capable of selective fluorescent turn-on recognition of the tumor biomarker α-fetoprotein. The technique is making use of amino-modified Mn-doped ZnS quantum dots (QDs) as solid supports, 4-vinylphenylboronic acid and methyl methacrylate as the functional monomers, γ-methacryloxypropyl trimethoxysilane as the grafting agent, and α-fetoprotein as a template. A graft imprint is created on the surface of the QDs. The functional monomers are shown to play an important role in the formation of the binding sites and in preventing nonspecific protein binding. The resulting MIP-QDs display a good linear response to α-fetoprotein in the 50 ng · L⁻¹ to 10 μg · L⁻¹ concentration range, and the limit of detection is 48 ng · L⁻¹. In our perception, the method has a wide scope in that it may be adapted to various other glycoproteins.

Schematic illustration of the synthesis of the MIP-QDs composites

     

Coaxial conducting polymer nanotubes: polypyrrole nanotubes coated with polyaniline or poly(<Emphasis Type="Italic">p</Emphasis>-phenylenediamine) and products of their carbonisation

Polypyrrole nanotubes were prepared by the oxidation of pyrrole with iron(III) chloride in a reaction mixture containing methyl orange. They were subsequently coated with polyaniline or poly(p-phenylenediamine) in situ during the oxidation of respective monomers in their presence. A part of the coaxial nanotubes was deprotonated using ammonia solution. The conductivity of polypyrrole nanotubes of 60 S cm^?1, was reduced after the coating, and again after the deprotonation, but maintained at a level above 10^?4 S cm^?1. Infrared and Raman spectra reflect the presence of the polymer overlayer deposited on the polypyrrole template. Thermogravimetric analysis was used as a tool for the analytical carbonisation of samples in an inert nitrogen atmosphere. The conversion of conducting polymers to nitrogen-containing carbon nanotubes was confirmed using Raman spectra.     

A Self-Assembled Unit Comprising 12 Squaraine Dyes Built Up from Two Star-Shaped Hexasquarainyl-Benzene Molecules

We describe the aggregate formation and optical properties of a star-shaped hexaarylbenzene with six squaraine chromophores (=hexasquarainyl benzene). Comprehensive concentration-dependence studies in acetone/CHCl₃ mixtures reveal a strong propensity to form discrete dimeric aggregates with a high binding constant in excess of 10⁶ m ⁻¹. In this context, a large hypsochromic shift of almost 2700 cm⁻¹ was found in the absorption spectrum, indicating H-type exciton coupling. The aggregate band is characterised by a very small band width of only 560 cm⁻¹, probably caused by exchange narrowing. Both experimental and computational methods were used to elucidate the supramolecular aggregate structure, which is assumed to consist of two stacked hexasquarainyl benzene monomers.     

Luminescence spectral properties of dendritic oligothiophenesilane macromolecules

The luminescence spectral properties of oligothiophenesilane dendrite macromolecules were studied. It was found that the chromophores responsible for the formation of the absorption and luminescence spectra were dendrimer fragments separated by silicon atoms; all the chromophores are equally involved in the formation of the absorption spectra of dendritic macromolecules of the zero, first, second, and third generations. It was shown that for dendrites of the first, second, and third generations, the luminescence spectrum is mainly formed by the chromophore fragments lying in the internal layers of the dendritic macromolecule due to the induction resonance energy transfer of electronic excitation from peripheral to internal chromophore fragments. The conclusion was drawn that synthesis of dendritic macromolecules with internal chromophore fragments possessing a high quantum yield of fluorescence can give actively luminescing nanosized objects with the molar extinction coefficient proportional to the number of fragments ɛ_max ≈ 10⁶ l/(mol cm). These compounds can find wide application in transducers for converting various types of ionizing radiation into optical radiation in electroluminescent devices.     

In‐Planta Expression: Searching for the Genuine Chromophores of Cryptochrome‐3 from Arabidopsis thaliana

Göbel et al. present in this issue an exemplary study of identification of chromophores from Arabidopsis thaliana cryptochrome‐3. Usually taken for granted, proteins and cofactors, respective chromophores, from heterologous expression are considered identical to material isolated from their genuine host. Cryptochromes carry two chromophores, an antenna cofactor and a functional flavin chromophore, both noncovalently embedded into the protein. In particular the antenna chromophore is loosely bound and often lost during protein purification. The authors identify from plant‐extracted Cry3 unambiguously N⁵,N¹⁰‐methenyltetrahydrofolate as antenna chromophore and flavin adenine dinucleotide as the functional chromophore.     

Direct differentiation of A‐ring single attachment versus A‐ and D‐ring double attachment of phycoerythrobilin chromophores to phycobiliproteins using MALDI mass spectrometry

Bilin chromophore attachment to phycobiliproteins is an enzyme‐catalyzed post‐translational modification process. Bilin‐lyases attach a bilin chromophore to their cognate protein through a thioether bond between the chromophore and a cysteine moiety. Bilin chromophores are attached to their phycobiliproteins through the 3¹ carbon of the bilin. Double attachment may also occur, and in this case, carbons 3¹ and 18¹ of the bilin are both forming covalent linkages to cysteine moieties. There is a mass spectrometric limitation when examining tryptic peptides containing two (or more) cysteines if one seeks to ascertain whether chromopeptides are singly or doubly attached. The problem is that singly and doubly attached chromopeptides appear at the same m/z value; thus, up until the present, only NMR analysis has been successful at determining whether the chromophore is singly or doubly attached. We report in this work a new, fast and accurate method for discriminating singly from doubly attached chromophores using MALDI‐TOF mass spectrometry. This method was developed from mass spectral analysis of chromopeptides that had undergone in vitro or in vivo attachment of bilin chromophores to phycobiliproteins. Distinction is based on a characteristic neutral loss that appears in the MALDI‐TOF mass spectrum only when the bilin is singly attached. Copyright © 2013 John Wiley & Sons, Ltd.     

N‐Annulated Perylene‐Substituted and Fused Porphyrin Dimers with Intense Near‐Infrared One‐Photon and Two‐Photon Absorption

Fusion of two N‐annulated perylene (NP) units with a fused porphyrin dimer along the S₀–S₁ electronic transition moment axis has resulted in new near‐infrared (NIR) dyes 1 a / 1 b with very intense absorption (ε>1.3×10⁵ M ^?1 cm^?1) beyond 1250 nm. Both compounds displayed moderate NIR fluorescence with fluorescence quantum yields of 4.4×10^?6 and 6.0×10^?6 for 1 a and 1 b , respectively. The NP‐substituted porphyrin dimers 2 a / 2 b have also been obtained by controlled oxidative coupling and cyclodehydrogenation, and they showed superimposed absorptions of the fused porphyrin dimer and the NP chromophore. The excited‐state dynamics of all of these compounds have been studied by femtosecond transient absorption measurements, which revealed porphyrin dimer‐like behaviour. These new chromophores also exhibited good nonlinear optical susceptibility with large two‐photon absorption cross‐sections in the NIR region due to extended π‐conjugation. Time‐dependent density functional theory calculations have been performed to aid our understanding of their electronic structures and absorption spectra.     

Darstellung,Eigenschaften und elektronische Raman-Spektren von Bis(chloro)phthalocyaninatoferrat(III), -ruthenat(III) und -osmat(III)

Preparation, Properties and Electronic Raman Spectra of Bis(chloro)-phthalocyaninatoferrate(III), -ruthenate(III) and -osmate(III) Bis(chloro)phthalocyaninatometalates of Fe^III, Ru^III and Os^III [MCl₂Pc(2-)]^?, with an electronic low spin ground state are formed by the reaction of [FeClPc(2-)] resp. H[MX₂Pc(2?)] (M = Ru, Os; X = Cl, I) with excess chloride in weakly coordinating solvents (DMF, THF) and are isolated as (n-Bu₄N) salts. The asym. M? Cl stretch (ν_as(MCl)) is observed in the f.i.r. at 288 cm^?1 (Fe), 295 cm^?1 (Ru), 298 cm^?1 (Os), ν_as(MN) at 330 cm^?1 (Fe), 327 cm^?1 (Ru), and 317 cm^?1 (Os); only ν_s(OsCl) at 311 cm^?1 is resonance Raman (r.r.) enhanced with blue excitation. The m.i.r. and FT-Raman spectra are typical for hexacoordinated phthalocyanines of tervalent metal ions. The UV-vis spectra show besides the characteristic π-π* transitions (B, Q, N, L band) of the Pc ligand a number of extra bands at 12–15 kK and 18–24 kK due to trip-doublet and (Pc→M)CT transitions. The effect of metal substitution is discussed. The r.r. spectra obtained by excitation between the B and Q band (λ₀ = 476.5 nm) are dominated by the intraconfigurational transition Γ₇ Γ ₈ arrising from the spin-orbit splitting of the electronic ground state for Fe^III at 536 cm^?1, for Ru^III at 961 cm^?1 and Os^III at 3 028 cm^?1. Thus the spin-orbit coupling constant increases very greatly down the iron group: Fe^III (357 cm^?1)< Ru^III (641 cm^?1)< Os^III (2 019 cm^?1). The Γ₇ Γ ₈-transition is followed by a very pronounced vibrational finestructure being composed in the r.r. spectra by the coupling with ν_s(MCl), δ(MClN) and the most intense fundamental vibrations of the Pc ligand. In absorption only vibronically induced transitions are observed for the Ru and Os complex at 1 700-2800 rsp. 3100-5800 em^?1 instead of the 0-0 phonon transitions. The most intense lines are attributed to combinations of the intense odd vibrational mo-des at ≈ 740 and 1120 cm^?1 with ν₅(MCI), δ(MClN).     

煤焦油的拉曼光谱表征和组分识别

用355 nm激光作为激发光源检测了煤焦油常温拉曼光谱;应用两种量子化学计算程序(Gaussian-DFT和ADF)模拟了占总量1%以上的15种煤焦油组分的拉曼光谱,模拟结果与实验光谱能较好匹配,并对振动模式进行了归属分析。研究表明,煤焦油组分主要由共轭六元环构成,其拉曼光谱特征谱带主要在1 660、1 420和1 265 cm^-1附近,当共轭六元环成链式结构时,1 420 cm^-1谱带特征明显;五元环嵌入共轭六元环链式结构会导致其拉曼光谱在1 265和1 660 cm^-1谱带相对强度增大;五元环、杂原子基团和甲基侧链依附在共轭六元环上,则对组分的拉曼光谱影响不显著。     

Proton magnetic resonance studies of compounds with bridgehead nitrogen atoms. IX Configurational and conformational studies with methyl-substituted 8-phenyl-l,8-diazabicyclo[4.3.0]nonanes

The synthesis of a series of methyl-substituted 8-phenyl-1,8-diazabicyclo[4.3.0]nonanes is described. The preferred conformations of these compounds have been assigned on the basis of the 2850-2500 cm^?1 region of their infrared spectra and of the geminal coupling constant of the C-9 methylene group.     

Propeller-Shaped Semi-fused Porphyrin Trimers: Molecular-Symmetry-Dependent Chiroptical Response

Triple helicene-like semi-fused trimeric Ni^II porphyrins were constructed by alkyne trimerization of an ethynyl-substituted porphyrin and subsequent three-fold Grignard addition to the formyl groups and acid-catalyzed intramolecular cyclization. The presence of stereogenic sp³ carbons in the central bridge leads to small inter-porphyrin conjugative interactions as was revealed by electrochemical and optical properties. Two diastereomers with stable chiral conformations were optically resolved, and the separated enantiomers displayed considerably intense circular dichroism. Importantly, the chiroptical response of C₃-symmetric helical isomer (|Δϵ|=830 m ⁻¹ cm⁻¹) is 1.8 times amplified from that of C₁-symmetric one (|Δϵ|=470 m ⁻¹ cm⁻¹). The observed amplification has been interpreted in terms of different spatial arrangements of the three porphyrins.     

The raman spectrum of biosynthetic human growth hormone. Its deconvolution,bandfitting, and interpretation

The Raman spectrum of amorphous biosynthetic human growth hormone, somatotropin, has been measured at high signal-to-noise ratios, using a CW argon ion laser and single channel detection. The rms signal-to-noise ratio varies from 1800:1 in the Amide I region near 1650 cm⁻¹ region, to 500:1 in the disulfide stretch region near 500 cm⁻¹.Component Raman bands have been extracted from the entire spectral envelope from 1800-400 cm⁻¹, by an interactive process involving both partial deconvolution and band-fitting. Interconsistency of all bands has been achieved by multiple overlapping of adjacent regions that had been isolated for the band-fitting programs.The resulting areas of the Raman component bands have been interpreted to show the ratios of peptide conformations in the hormone: 64% α-helix, 24% β-sheet, 8% β-turns and 4% γ-turns. Analysis of the tyrosine region, usually described as a Fermi resonance doublet near ∼830–850 cm⁻¹, shows four bands, at 825, 833, 853, and 859 cm⁻¹ in this macromolecule. Integrated intensities of these bands (2:2:2:2) are interpreted to show that only half of the eight tyrosine residues function as hydrogen-bond bridges via the acceptance of protons.Both disulfide bridges fall within the frequency ranges for normal, unstressed SS bonds: The 511 and 529 cm⁻¹ bands are indicative of the gauche-gauche-gauche and trans-gauche-gauche conformations, respectively.     

ENERGY TRANSFER STUDIES ON CRYPTOMONAD BILIPROTEINS

Abstract. Fluorescence techniques of various types have been used to study the light-gathering and energy transfer modes for various cryptomonad biliproteins (phycocyanin or phycoerythrins). Analysis of fluorescence polarization and absorption data demonstrates that each cryptomonad biliprotein is composed of at least two distinct types of absorbing chromophore, each attached to the protein through covalent linkages to different polypeptide chains. Examination of the fluorescence emission spectra as a function of excitation at several wavelengths demonstrates that only one of these absorbing chromo-phores is responsible for the fluorescence. This behavior is consistent with a known phenomenon whereby photons are gathered by more than one chromophore and then after radiationless energy transfer are emitted by only one chromophore. Application of Förster dipole-dipole energy transfer theory is made to the study of the mode by which energy absorbed by biliproteins migrates to Chl a. The spectral overlap integral between phycocyanin (Chroomonas sp.) and Chl a is 7.13 ± 10^-10cm⁶mol^-1and between phycocyanin and Chl c₂ 0.25 ± 10^-10cm⁶mol^-1. This large difference in overlap suggests, although does not prove, that phycocyanin might transfer energy directly to Chl a without a Chl c₂ intermediary. The cryptomonad phycoerythrins may also use this method but a Chl c₂ intermediate could not be ruled out for them. Radiationless energy transfer among homogeneous biliproteins is shown to be feasible. All these calculations are based on in vitro spectra and the interpretations extrapolated to the cellular situation, and these tentative conclusions are reached without knowledge of other factors, such as chromophore-chro-mophore orientation and distance, which could greatly influence the energy transfer scheme.     

ULTRAVIOLET ERYTHEMA SENSITIVITY IN ANAMNESTIC (I-IV) and PHOTOTESTED (1–4) CAUCASIAN SKIN PHOTOTYPES: THE NEED FOR A NEW CLASSIFICATION SYSTEM

Abstract— The anamnestic skin phototypes (ASP) I-IV^1,2 of 21 Caucasian volunteers were compared with their phototested skin phototypes (PSP) using solar simulating, broadband UV radiation. The Commission Internationale de' éclairage (CIE)-weighted (i.e. erythemally effective) minimal erythema doses (MED) for solar simulating radiation varied from 20 mJ/cm² (PSP type 1) to 57 mJ/cm² (PSP type 4). In only 11 of 21 volunteers did the ASP (I-IV) and PSP (1–4) classifications coincide, and the MED values of the volunteers within the different ASP groups (I-IV) overlapped considerably. To compare the reactivity to erythematogenic radiation of different wavelengths, narrowband monochromator irradiations were performed at 298 nm, 310 nm and 330 nm. The CIE-weighted MED values at these wavelengths (20–80 mJ/cm²) corresponded well with those obtained in the broadband testing. Our results indicate that, with classification by interrogation, Caucasian skin can reliably be classified into only two subtypes, corresponding to Fitzpatrick phototypes I–III and phototype IV, respectively. A classification into four sensitivity types can be achieved by phototesting, only. We propose that the concept of ASP should be used with caution. The concept of PSP 1–4 should be favored.     

A high pressure spectroscopic study on the ice III — ice IX. Disordered — ordered transition

The only published data on the disordered-to-ordered ice III to ice IX transition refers to measurements of di-electric constant. Raman spectra of ice III and ice IX were recorded under a pressure of 0.3 GPa for temperatures in the range 250 to 130 K. They clearly show a transition that is predominantly of the disordered-ordered type. Raman spectra in the frequency range 15–4000 cm^?1 will be shown but special attention will be given to two translational lattice modes at about 190 cm^?1 and 65.5 cm^?1 which show somewhat unusual behaviour. Small discontinuities in the frequency versus temperature plots suggest that there is a small discontinuous decrease in the volume during the ice III to ice IX transition.