Vibrational Lifetime of the SCN Protein Label in H2O and D2O Reports Site-Specific Solvation and Structure Changes During PYP's Photocycle.

The application of vibrational labels such as thiocyanate  (-S-C≡N) for studying protein structure and dynamics is thriving. Absorption spectroscopy is usually employed to obtain wavenumber and line shape of the label. An observable of great significance might be the vibrational lifetime, which can be obtained by pump probe or 2D-IR spectroscopy. Due to the insulating effect of the heavy sulfur atom in the case of the SCN label, the lifetime of the C≡N oscillator is expected to be particularly sensitive to its surrounding as it is not dominated by through-bond relaxation. We therefore investigate the vibrational lifetime of the SCN label at various positions in the blue light sensor protein Photoactive Yellow Protein (PYP) in the ground state and signaling state of the photoreceptor. We find that the vibrational lifetime of the C≡N stretching mode is strongly affected both by its protein environment and by the degree of exposure to the solvent. Even for label positions where the line shape and wavenumber observed by FTIR are barely changing upon activation of the photoreceptor, we find that the lifetime can change considerably. To obtain an unambiguous measure for the solvent exposure of the labeled site, we show that it is imperative to compare the lifetimes in H2O and D2O. Importantly, the lifetimes shorten in H2O as compared to D2O for water exposed labels, while they stay largely the same for buried labels. We quantify this effect by defining a solvent exclusion coefficient (SEC). The response of the label's vibrational lifetime to its solvent exposure renders it a suitable universal probe for protein investigations. This applies even to systems that are otherwise hard to address, such as transient or short-lived states, which could be created during a protein's working cycle (as here in PYP) or during protein folding. It is also applicable to flexible systems (intrinsically disordered proteins), protein-protein and protein-membrane interactions.

Ultrasound-assisted thiocyanation of aromatic and heteroaromatic compounds using ammonium thiocyanate and DDQ.

Thiocyanation of various aromatic and heteroaromatic compounds has been achieved in the presence of ammonium thiocyanate (NH(4)SCN) and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) in methanol solution under ultrasound irradiation. An ultrasonic probe of 24 kHz frequency has been used for this study. Whereas the use of ultrasound increases the rate of reactions compared with reactions at reflux conditions, the electron-donor ability of aromatic nucleus enhances also the rate of reaction.

[Radiolysis of aqueous thiocyanate and its complicated system].

We studied the 60Co gamma-ray radiolysis of thiocyanate aqueous solutions in the presence of free radical scavengers, determination of cyanide of radiated thiocyanate solutions, degradation of cyanide and thiocyanate complicated system and cyanide, thiocyanate and copper (I) cyanide complicated system. The presence of NaHCO3 and n-butanol made the degradation efficiency of thiocyanate decreased, in the radiation procession of thiocyanate, cyanide was produced; and the possible reaction equation is SCN(-) + 8*OH-->SO4(2-) + CN(-) +4H2O, in the cyanide and thiocyanate system, the thiocyanate degradation rate can obtained 30%, free cyanide degradation was little effected by coexistence of thiocyanate; and due to strong complex ability of copper (I), the main form of copper (I) cyanide is Cu(CN)3(2-), the total cyanide of the system can't be degraded completely and degradation rate is about 80%.

Photocurrent-voltage of a dye-sensitized nanocrystalline TiO2 solar cells influenced by N719 dye adsorption properties.

Titanium particles of single-phase anatase nanocrystallites were prepared by the hydrolysis of titanium tetraisopropoxide. A dye-sensitized solar cell (DSSC) was fabricated by adsorbing cis-bis(isothiocyanato)bis(2,2'-bipyridyl-4,4'-dicarboxylato)-ruthenium(II)bis-tetrabutylammonium dye (N719) onto TiO2 film. The samples were characterized by XRD, TEM, FE-SEM, AFM, and Brunauer-Emmett-Teller (BET) analysis. The influence of the acetic acid treatment of TiO2 electrode with different concentrations on the photovoltaic performance of DSSC was investigated. It was found that DSSC had better photoelectric performance when the TiO2 electrode was treated by acetic acid of 0.5 M. An equivalent circuit analysis using the one-diode model was used to evaluate the influences of adsorption quantity and acetic acid treatment on the energy conversion efficiency of DSSC. A nonlinear least-square optimization method was used to determine five model parameters.

Digital imaging fluorescence microscopy: spatial heterogeneity of photobleaching rate constants in individual cells.

Photobleaching and related photochemical processes are recognized experimental barriers to quantification of fluorescence by microscopy. We have measured the kinetics of photobleaching of fluorophores in living and fixed cells and in microemulsions, and have demonstrated the spatial variability of these processes within individual cells. An inverted fluorescence microscope and a high-sensitivity camera, together with high-speed data acquisition by a computer-controlled image processor, have been used to control precisely exposure time to excitation light and to record images. To improve the signal-to-noise ratio, 32 digital images were integrated. After correction for spatial variations in camera sensitivity and background fluorescence, the images of the relative fluorescence intensities for 0.065 micron2 areas in the object plane were obtained. To evaluate photobleaching objectively, an algorithm was developed to fit a three-parameter exponential equation to 20 images recorded from the same microscope field as a function of illumination time. The results of this analysis demonstrated that the photobleaching process followed first-order reaction kinetics with rate constants that were spatially heterogeneous and varied, within the same cell, between 2- and 65-fold, depending on the fluorophore. The photobleaching rate constants increased proportionally with increasing excitation intensity and, for benzo(a)pyrene, were independent of probe concentration over three orders of magnitude (1.25 microM to 1.25 mM). The propensity to photobleach was different with each fluorophore. Under the cellular conditions used in these studies, the average rates of photobleaching decreased in this order: N-(7-nitrobenz-2-oxa-1,3-diazole)-23,24-dinor-5-cholen-22-amine-3 beta-ol greater than acridine orange greater than rhodamine-123 greater than benzo(a)pyrene greater than fluorescein greater than tetramethylrhodamine greater than 1,1'dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine. The photobleaching appears to be an oxidation reaction, in that the addition of saturated solutions of Na2S2O5 to mineral oil microemulsions eliminated photobleaching of N-(7-nitrobenz-2-oxa-1,3-diazole)-23,24-dinor-5-cholen-22-amine-3 beta-ol or benzo(a)pyrene. We identified experimental conditions to observe, without detectable photobleaching, fluorophores in living cells, which can not be studied anaerobically. Useful images were obtained when excitation light was reduced to eliminate photobleaching, as determined from zero-time images calculated from the exponential fit routine.(ABSTRACT TRUNCATED AT 400 WORDS)