Thermodynamic analysis of a molecular chaperone binding to unfolded protein substrates.

Molecular chaperones are a highly diverse group of proteins that recognize and bind unfolded proteins to facilitate protein folding and prevent nonspecific protein aggregation. The mechanisms by which chaperones bind their protein substrates have been studied for decades. However, there are few reports about the affinity of molecular chaperones for their unfolded protein substrates. Thus, little is known about the relative binding affinities of different chaperones and about the relative binding affinities of chaperones for different unfolded protein substrates. Here we describe the application of SUPREX (stability of unpurified proteins from rates of H-D exchange), an H-D exchange and MALDI-based technique, in studying the binding interaction between the molecular chaperone Hsp33 and four different unfolded protein substrates, including citrate synthase, lactate dehydrogenase, malate dehydrogenase, and aldolase. The results of our studies suggest that the cooperativity of the Hsp33 folding-unfolding reaction increases upon binding with denatured protein substrates. This is consistent with the burial of significant hydrophobic surface area in Hsp33 when it interacts with its substrate proteins. The SUPREX-derived K(d) values for Hsp33 complexes with four different substrates were all found to be within the range of 3-300 nM.

A low-cost, portable, and quantitative spectral imaging system for application to biological tissues.

The ability of diffuse reflectance spectroscopy to extract quantitative biological composition of tissues has been used to discern tissue types in both pre-clinical and clinical cancer studies. Typically, diffuse reflectance spectroscopy systems are designed for single-point measurements. Clinically, an imaging system would provide valuable spatial information on tissue composition. While it is feasible to build a multiplexed fiber-optic probe based spectral imaging system, these systems suffer from drawbacks with respect to cost and size. To address these we developed a compact and low cost system using a broadband light source with an 8-slot filter wheel for illumination and silicon photodiodes for detection. The spectral imaging system was tested on a set of tissue mimicking liquid phantoms which yielded an optical property extraction accuracy of 6.40 +/- 7.78% for the absorption coefficient (micro(a)) and 11.37 +/- 19.62% for the wavelength-averaged reduced scattering coefficient (micro(s)').

Supercontinuum generation in air-silica microstructured fibers with nanosecond and femtosecond pulse pumping

We study the generation of supercontinua in air-silica microstructured fibers by both nanosecond and femtosecond pulse excitation. In the nanosecond experiments, a 300-nm broadband visible continuum was generated in a 1.8-m length of fiber pumped at 532 nm by 0.8-ns pulses from a frequency-doubled passively Q-switched Nd:YAG microchip laser. At this wavelength, the dominant mode excited under the conditions of continuum generation is the LP 11 mode, and, with nanosecond pumping, self-phase modulation is negligible and the continuum generation is dominated by the interplay of Raman and parametric effects. The spectral extent of the continuum is well explained by calculations of the parametric gain curves for four-wave mixing about the zero-dispersion wavelength of the LP11 mode. In the femtosecond experiments, an 800-nm broad-band visible and near-infrared continuum has been generated in a 1-m length of fiber pumped at 780 nm by 100-fs pulses from a Kerr-lens model-locked Ti:sapphire laser. At this wavelength, excitation and continuum generation occur in the LP01 mode, and the spectral width of the observed continuum is shown to be consistent with the phase-matching bandwidth for parametric processes calculated for this fiber mode. In addition, numerical simulations based on an extended nonlinear Schrödinger equation were used to model supercontinuum generation in the femtosecond regime, with the simulation results reproducing the major features of the experimentally observed spectrum. © 2002 Optical Society of America.

Mathematical modelling: a laser soldering process for an optoelectronics butterfly package

For sensitive optoelectronic components, traditional soldering techniques cannot be used because of their inherent sensitivity to thermal stresses. One such component is the Optoelectronic Butterfly Package which houses a laser diode chip aligned to a fibre-optic cable. Even sub-micron misalignment of the fibre optic and laser diode chip can significantly reduce the performance of the device. The high cost of each unit requires that the number of damaged components, via the laser soldering process, are kept to a minimum. Mathematical modelling is undertaken to better understand the laser soldering process and to optimize operational parameters such as solder paste volume, copper pad dimensions, laser solder times for each joint, laser intensity and absorption coefficient. Validation of the model against experimental data will be completed, and will lead to an optimization of the assembly process, through an iterative modelling cycle. This will ultimately reduce costs, improve the process development time and increase consistency in the laser soldering process.

Low temperature crystal structures of two rhodanine derivatives, 3-amino rhodanine and 3-methyl rhodanine: geometry of the rhodanine ring

Rhodanines (2-thio-4-oxothiazolidines) are synthetic small molecular weight organic molecules with diverse applications in biochemistry, medicinal chemistry, photochemistry, coordination chemistry and industry. The X-ray crystal structure determination of two rhodanine derivatives, namely (I), 3-aminorhodanine [3-amino-2-thio-4-oxothiazolidine], C3H4N2OS2, and (II) 3-methylrhodanine [3-methyl-2-thio-4-oxothiazolidine], C4H5NOS2, have been conducted at 100 K. I crystallizes in the monoclinic space group P2(1)/n with unit cell parameters a = 9.662(2), b = 9.234(2), c = 13.384(2) angstrom, beta = 105.425(3)degrees, V = 1151.1(3) angstrom(3), Z = 8 (2 independent molecules per asymmetric unit), density (calculated) = 1.710 mg/m(3), absorption coefficient = 0.815 mm(-1). II crystallizes in the orthorhombic space group Iba2 with unit cell a = 20.117(4), b = 23.449(5), c = 7.852(2) angstrom, V = 3703.9(12) angstrom(3), Z = 24 (three independent molecules per asymmetric unit), density (calculated) = 1.584 mg/m(3), absorption coefficient 0.755 mm(-1). For I in the final refinement cycle the data/restraints/parameter ratios were 2639/0/161, goodness-of-fit on F-2 = 0.934, final R indices [I > 2sigma(I)] were R1 = 0.0299, wR2 = 0.0545 and R indices (all data) R1 = 0.0399, wR2 = 0.0568. The largest difference peak and hole were 0.402 and -0.259 e angstrom(-3). For II in the final refinement cycle the data/restraints/parameter ratios were 3372/1/221, goodness-of-fit on F(2) = 0.950, final R indices [I > 2sigma(I)] were R1 = 0.0407, wR2 = 0.1048 and R indices (all data) R1 = 0.0450, wR2 = 0.1088. The absolute structure parameter = 0.19(9) and largest difference peak and hole 0.934 and -0.301 e angstrom(-3). Details of the geometry of the five molecules (two for I and three for II) and the crystal structures are fully discussed. Corresponding features of the molecular geometry are highly consistent and firmly establish the geometry of the rhodanine

Low temperature crystal structure of N-Acetyl-L-Glutamic acid: comparison with the DFT calculated structure

N-acetyl-L-glutamic acid, crystallizes in the orthorhombic space group P2(1)2(1)2(1) with unit cell parameters a = 4.747(3), b = 12.852(7), c = 13.906(7) Å, V = 848.5(8) Å3, Z = 4, density (calculated) = 1.481 mg/m3, linear absorption coefficient 0.127 mm−1. The crystal structure determination was carried out with MoKalpha X-ray data measured with liquid nitrogen cooling at 100(2) K temperature. In the final refinement cycle the data/restraints/parameter ratios were 1,691/0/131; goodness-of-fit on F(2) = 1.122. Final R indices for [I > 2sigma(I)] were R1 = 0.0430, wR2 = 0.0878 and R indices (all data) R1 = 0.0473, wR2 = 0.0894. The largest electron density difference peak and hole were 0.207 and −0.154 eÅ(−3). Details of the molecular geometry are discussed and compared with a model DFT structure calculated using Gaussian 98.

Oxygen photolysis in the Mauritanian upwelling: Implications for net community production

We carried out 16 photochemical experiments of filtered surface water in a custom-built solar simulator and concomitant measurements of in vitro gross primary production (GPP) and respiration (R) in the Mauritanian upwelling during a Lagrangian study following three sulfur hexafluoride–labeled patches of upwelled water (P1 to P3). Oxygen photolysis rates were correlated with the absorbance of chromophoric dissolved organic matter (CDOM) at 300 nm, suggesting first-order kinetics with respect to CDOM. An exponential fit was used to calculate the apparent quantum yield (AQY) for oxygen photolysis, giving an average AQY of 0.00053 µmol O2 (mole photons m−2 s−1)−1 at 280 nm and slope of 0.0012 nm−1. Modeled photochemical oxygen demand (POD) at the surface (3–16 mmol m−3 d−1) occasionally exceeded R and was dominated by ultraviolet radiation (71–79%). Euphotic-layer integrated GPP decreased with time during both P-1 and P-3, whereas R remained relatively constant and POD increased during P-1 and decreased during P-3. On Day 4 of P-3, GPP and POD maxima coincided with high CDOM absorbance, suggesting “new” CDOM production. Omitting POD may lead to an underestimation of net community production (NCP), both through in vitro and geochemical methods (here by 2–22%). We propose that oxygen-based NCP estimates should be revised upward. For the Mauritanian upwelling, the POD-corrected NCP was strongly correlated with standard NCP with a slope of 1.0066 ± 0.0244 and intercept of 46.51 ± 13.15 mmol m−2 d−1.

Toward autonomous measurements of photosynthetic electron transport rates: An evaluation of active fluorescence-based measurements of photochemistry

This study presents a methods evaluation and intercalibration of active fluorescence-based measurements of the quantum yield ( inline image) and absorption coefficient ( inline image) of photosystem II (PSII) photochemistry. Measurements of inline image, inline image, and irradiance (E) can be scaled to derive photosynthetic electron transport rates ( inline image), the process that fuels phytoplankton carbon fixation and growth. Bio-optical estimates of inline image and inline image were evaluated using 10 phytoplankton cultures across different pigment groups with varying bio-optical absorption characteristics on six different fast-repetition rate fluorometers that span two different manufacturers and four different models. Culture measurements of inline image and the effective absorption cross section of PSII photochemistry ( inline image, a constituent of inline image) showed a high degree of correspondence across instruments, although some instrument-specific biases are identified. A range of approaches have been used in the literature to estimate inline image and are evaluated here. With the exception of ex situ inline image estimates from paired inline image and PSII reaction center concentration ( inline image) measurements, the accuracy and precision of in situ inline image methodologies are largely determined by the variance of method-specific coefficients. The accuracy and precision of these coefficients are evaluated, compared to literature data, and discussed within a framework of autonomous inline image measurements. This study supports the application of an instrument-specific calibration coefficient ( inline image) that scales minimum fluorescence in the dark ( inline image) to inline image as both the most accurate in situ measurement of inline image, and the methodology best suited for highly resolved autonomous inline image measurements.

Restricted rotation dynamics and far-infrared spectra of liquid water governed by elastic interactions

A semi-phenomenological model describing wideband dielectric and far-infrared spectra of liquid water was proposed recently by the same authors [J. Mol. Struct. 606 (2002) 9], where a small dipole-moment component changing harmonically with time determines a weak absorption band (termed here the R-band) centred at the wavenumber v similar to 200 cm(-1). In the present work, a rough molecular theory of the R-band based on the concept of elastic interactions is given. Stretching and bending of hydrogen bonds cause restricted rotation (RR) of a polar water molecule in terms of a dimer comprising the H- bonded molecules. Analytical expression for the RR frequency nu(str) is derived as a function of the RR amplitude, geometrical parameters and force constants. The density g(nu(str)) of frequency distribution is shown to be centred in the R-band. The spectrum of the dipolar auto-correlation function calculated for this structural-dynamical model is found. A composite model comprising two intermolecular potentials is proposed, which yields for water a good description of the experimental wideband (from 0 to 1000 cm(- 1)) spectra of complex permittivity and of absorption coefficient. The presented interpretation of these spectra is based on a concept that water presents a two-component solution, with components differing by the types of molecular rotation. (C) 2003 Elsevier B.V. All rights reserved.

Specific and unspecific interactions in polar fluids in view of wideband dielectric far-infrared spectra

A simple molecular analytical theory of dielectric relaxation in strongly polar fluids is considered in terms of a semi- phenomenological approach. Theoretical spectra epsilon(v), a(v) of complex permittivity and absorption coefficient are fully determined by a form of intermolecular potential well, in which a dipole reorients. In a recent publication by VI. Gaiduk, O.F. Nielsen, and T.S. Perova [J. Molliq 95 (1002) 1-25] the wideband spectra of liquid H2O and D2O were described in terms of a composite model comprising the rectangular and the cosine squared potential wells. Much better results are achieved in this work, where the rectangular well is replaced by a well with a rounded bottom termed the hat-curved well. The spectrum of the auto-correlation function (ACF) is calculated for such a potential. The proposed theory of a composite model, comprising hat-curved and parabolic wells, is applied for liquid water. This model is capable for describing the Debye relaxation region, the second relaxation region in the submillimeter wavelength range, and the far infra-red (FIR) e(v), a(v) spectra, where an intense librational band and an additional weak band are placed, respectively, near 700 cm(-1) and 200 cm(-1). The latter band reflects the features of so-called specific (viz. directly related to H-bonds) interactions and the former band reflects the features of unspecific interactions. The physical mechanisms connected with these types of interactions are discussed in terms of two relevant types of water structure (types of molecular rotation). The proposed theory is also applied to a non-associated liquid in terms of one hat-curved potential well. (C) 2004 Elsevier B.V. All rights reserved.

A Dual Role for Lecithin:Cholesterol Acyltransferase (EC 2.3.1.43) in Lipoprotein Oxidation

Lecithin:cholesterol acyltransferase (LCAT) is a key enzyme involved in lipoprotein metabolism. It mediates the transesterification of free cholesterol to cholesteryl ester in an apoprotein A-I-dependent process. We have isolated purified LCAT from human plasma using anion-exchange chromatography and characterized the extracted LCAT in terms of its molecular weight, molar absorption coefficient, and enzymatic activity. The participation of LCAT in the oxidation of very low density lipoproteins (VLDL) and low-density lipoproteins (LDL) was examined by supplementing lipoproteins with exogenous LCAT over a range of protein concentrations. LCAT-depleted lipoproteins were also prepared and their oxidation kinetics examined. Our results provide evidence for a dual role for LCAT in lipoprotein oxidation, whereby it acts in a dose-responsive manner as a potent pro-oxidant during VLDL oxidation, but as an antioxidant during LDL oxidation. We believe this novel pro-oxidant effect may be attributable to the LCAT-mediated formation of oxidized cholesteryl ester in VLDL, whereas the antioxidant effect is similar to that of chain-breaking antioxidants. Thus, we have demonstrated that the high-density lipoprotein-associated enzyme LCAT may have a significant role to play in lipoprotein modification and hence atherogenesis. (C) 2007 Elsevier Inc. All rights reserved.

Control of the thickness of mesoporous titania films for application in multiphase catalytic microreactors

A new method of sol-gel polymer template synthesis of mesoporous catalytic thin films has been proposed which allows controlling the chemical nature of the film, the porosity, thickness and loading with an active species. The mesoporous films with a long-order structure can be obtained in a narrow range of surfactant-to-metal precursor molar ratios from 0.006 to 0.009. The catalytic film thickness was varied from 300 to 1000 nm while providing a uniform catalyst distribution with a desired catalyst loading (1 wt. % Au nanoparticles) throughout the film. The films were characterized by TEM, SEM, ethanol adsorption and contact angle measurements. The calcination of the as-synthesized films at 573 K reduced Ti4+ sites to Ti3+. A 300 nm thick Au-containing film showed an initial TOF of 1.4 s(-1) and a selectivity towards unsaturated alcohols as high as 90% in the hydrogenation of citral. Thicker films demonstrated a high selectivity towards the saturated aldehyde (above 55%) and a lower intrinsic catalytic activity (initial TOF of 0.7-0.9 s(-1)) in the absence of internal diffusion limitations.

Energy Dependence of Gold Nanoparticle Radiosensitization in Plasmid DNA

Gold nanoparticles (GNPs) are of considerable interest for use as a radiosensitizer, because of their biocompatibility and their ability to increase dose deposited because of their high mass energy absorption coefficient. Their sensitizing properties have been verified experimentally, but a discrepancy between the experimental results and theoretical predictions suggests that the sensitizing effect does not depend solely on gold's superior absorption of energetic photons. This work presents the results of three sets of experiments that independently mapped out the energy dependence of the radiosensitizing effects of GNPs on plasmid DNA suspended in water. Incident photon energy was varied from 11.8 to 80 keV through the use of monochromatic synchrotron and broadband X-rays. These results depart significantly from the theoretical predictions in two ways: First, the sensitization is significantly larger than would be predicted; second, it does not vary with energy as would be predicted from energy absorption coefficients. These results clearly demonstrate that the effects of GNP-enhanced therapies cannot be predicted by considering additional dose alone and that a greater understanding of the processes involved is necessary for the development of future therapeutics.

Nanodosimetric effects of gold nanoparticles in megavoltage radiation therapy

Background and purpose: The addition of gold nanoparticles (GNPs) to tumours leads to an increase in dose due to their high density and energy absorption coefficient, making it a potential radiosensitiser. However, experiments have observed radiosensitisations significantly larger than the increase in dose alone, including at megavoltage energies where gold's relative energy absorption is lowest. This work investigates whether GNPs create dose inhomogeneities on a sub-cellular scale which combine with non-linear dose dependence of cell survival to be the source of radiosensitisation at megavoltage energies.

Physical basis and biological mechanisms of gold nanoparticle radiosensitization

The unique properties of nanomaterials, in particular gold nanoparticles (GNPs) have applications for a wide range of biomedical applications. GNPs have been proposed as novel radiosensitizing agents due to their strong photoelectric absorption coefficient. Experimental evidence supporting the application of GNPs as radiosensitizing agents has been provided from extensive in vitro investigation and a relatively limited number of in vivo studies. Whilst these studies provide experimental evidence for the use of GNPs in combination with ionising radiation, there is an apparent disparity between the observed experimental findings and the level of radiosensitization predicted by mass energy absorption and GNP concentration. This review summarises experimental findings and attempts to highlight potential underlying biological mechanisms of response in GNP radiosensitization.