Influence of phosphate ligands in abolishing the conformational difference between ribonuclease A and its acid-denatured derivative

The initial structural alteration of RNAase A due to acid denaturation (0.5 N HCl, 30 degrees C) that accompanies deamidation (without altering enzymic activity) has been dectected by spectrophotometric titration, fluorescence and ORD/CD measurements. It is shown that acid treated RNAase A has an altered conformation at neutral pH, 25 degrees C. This is characterized by the increased accessibility of buried tyrosine residue(s) towards the solvent. The most altered conformation of RNAase A is found in the 10 h acid-treated derivative. This has about 1.5 additional exposed tyrosine residues and a lesser amount of secondary structure than RNAase A. All three methods (titration, fluorescence and CD) established that the structural transition of RNAase A is biphasic. The first phase occurs within 1 h and the resulting subtle conformational change is constant up to 7 h. Following this, after the release of 0.55 mol of ammonia, the major conformational change begins. The altered conformation of the acid-denatured RNAase A could be reversed completely to the native state through a conformational change induced by substrate analogs like 2'- or 3'-CMP. Thus the monodeamidated derivative isolated from the acid-denatured RNAase A by phosphate is very similar to RNAase A in over-all conformation. The results suggest the possibility of flexibility in the RNAase A molecule that does not affect its catalytic activity, as probed through the tyrosine residues.

Droplet Epitaxy of InN Quantum Dots on Si(111) by RF Plasma-Assisted Molecular Beam Epitaxy

InN quantum dots (QDs) were fabricated on Si(111) substrate by droplet epitaxy using an RF plasma-assisted MBE system. Variation of the growth parameters, such as growth temperature and deposition time, allowed us to control the characteristic size and density of the QDs. As the growth temperature was increased from 100 C to 300 degrees C, an enlargement of QD size and a drop in dot density were observed, which was led by the limitation of surface diffusion of adatoms with the limited thermal energy. Atomic force microscopy (AFM) and scanning electron microscopy (SEM) were used to assess the QDs size and density. The chemical bonding configurations of InN QDs were examined by X-ray photo-electron spectroscopy (XPS). Fourier transform infrared (FTIR) spectrum of the deposited InN QDs shows the presence of In-N bond. Temperature-dependent photoluminescence (PL) measurements showed that the emission peak energies of the InN QDs are sensitive to temperature and show a strong peak emission at 0.79 eV.

Diffusion coefficients for selected binary liquid systems: cyclohexanes and n-alkyl alcohols

In the present work the integral diffusion coefficients are estimated by using the diaphragm cell technique. The diffusion coefficients are measured at various compositions for two sets binary systems: one of cyclohexane and n-paraffinic alcohols and the other of methylcyclohexane and n-paraffinic alcohols. The alcohols used are seven members of homologous series of n-paraffinic alcohols from ethanol to octanol. The maximum possible error in the experimental diffusion coefficient could be 8% for both the cyclohexane-n-alkyl alcohol system and methylcyclohexane-n-alkyl alcohol system. A correlation for each of the two sets of binary systems is given. The maximum deviation in the correlations was less than 6.5 and 3.5% for cyclohexane-n-alkyl alcohols and methylcyclohexane-n-alkyl alcohols, respectively.

Optimization of two-dimensional NMR by matched accumulation

It is well known that in the time-domain acquisition of NMR data, signal-to-noise (S/N) improves as the square root of the number of transients accumulated. However, the amplitude of the measured signal varies during the time of detection, having a functional form dependent on the coherence detected. Matching the time spent signal averaging to the expected amplitude of the signal observed should also improve the detected signal-to-noise. Following this reasoning, Barna et al. (J Magn. Reson.75, 384, 1987) demonstrated the utility of exponential sampling in one- and two-dimensional NMR, using maximum-entropy methods to analyze the data. It is proposed here that for two-dimensional experiments the exponential sampling be replaced by exponential averaging. The data thus collected can be analyzed by standard fast-Fourier-transform routines. We demonstrate the utility of exponential averaging in 2D NOESY spectra of the protein ubiquitin, in which an enhanced SIN is observed. It is also shown that the method acquires delayed double-quantum-filtered COSY without phase distortion.

Visualisation of a 2′-5′ parallel stranded double helix at atomic resolution: crystal structure of cytidylyl-2′,5′-adenosine

X-ray crystallographlc studies on 3′–5′ ollgomers have provided a great deal of information on the stereochemistry and conformational flexibility of nucleic acids and polynucleotides. In contrast, there is very little Information available on 2′–5′ polynucleotides. We have now obtained the crystal structure of Cytidylyl-2′,5′-Adenoslne (C2′p5′A) at atomic resolution to establish the conformational differences between these two classes of polymers. The dlnucleoside phosphate crystallises in the monocllnlc space group C2, with a = 33.912(4)Å, b =16.824(4)Å, c = 12.898(2)Å and 0 = 112.35(1) with two molecules in the asymmetric unit. Spectacularly, the two independent C2′p5′A molecules in the asymmetric unit form right handed miniature parallel stranded double helices with their respective crystallographic two fold (b axis) symmetry mates. Remarkably, the two mini duplexes are almost indistinguishable. The cytosines and adenines form self-pairs with three and two hydrogen bonds respectively. The conformation of the C and A residues about the glycosyl bond is anti same as in the 3′–5′ analog but contrasts the anti and syn geometry of C and A residues in A2′p5′C. The furanose ring conformation is C3′endo, C2′endo mixed puckering as in the C3′p5′A-proflavine complex. A comparison of the backbone torsion angles with other 2′–5′ dinucleoside structures reveals that the major deviations occur in the torsion angles about the C3′–C2′ and C4′-C3′ bonds. A right-handed 2′–5′ parallel stranded double helix having eight base pairs per turn and 45° turn angle between them has been constructed using this dinucleoside phosphate as repeat unit. A discussion on 2′–5′ parallel stranded double helix and its relevance to biological systems is presented.

Kinetics of propellant decomposition

Equations for solid-state decompositions which are controlled by the phase-boundary movement and nucleation have been examined using ammonium perchlorate/polystyrene propellant decomposition at 503 K and 533 K. It was found that 3 different equations governed by the nucleation process show a good fit of data at these temperatures. However, the best fit was obtained for the following Avrami-Erofeev equation, [-In (1 - α]1/4=kt.

Interaction of rhodium(III) with DNA

The nature of interaction of Rh(III) with DNA was studied using viscometry and ultraviolet, visible and infrared spectroscopy. The rate of interaction was found to be very slow at room temperature taking several days for completion. The time needed to attain equilibrium is dependent on the concentrations of metal ion, higher the concentration shorter the period required for equilibration. Visible spectra of Rh(III) were found to alter considerably in the presence of DNA. An increase in absorbance and a red shift were observed in the ultraviolet spectra of DNA in the presence of Rh(III). The specific viscosity of DNA solution was found to decrease asymptotically with time and concentrations of metal ion. The melting temperature of DNA was found to increase at lower metal ion concentrations, whereas at higher values a decrease was obtained. At still higher metal ion concentrations (Image ) a ‘nonmeltable state’ of DNA was observed. These results seem to indicate that Rh(III) binds both with the phosphate and the bases of the DNA.

Analytical and numerical solutions of inward spherical solidification of a superheated melt with radiative-convective heat transfer and density jump at freezing front

Analytical and numerical solutions of a general problem related to the radially symmetric inward spherical solidification of a superheated melt have been studied in this paper. In the radiation-convection type boundary conditions, the heat transfer coefficient has been taken as time dependent which could be infinite, at time,t=0. This is necessary, for the initiation of instantaneous solidification of superheated melt, over its surface. The analytical solution consists of employing suitable fictitious initial temperatures and fictitious extensions of the original region occupied by the melt. The numerical solution consists of finite difference scheme in which the grid points move with the freezing front. The numerical scheme can handle with ease the density changes in the solid and liquid states and the shrinkage or expansions of volumes due to density changes. In the numerical results, obtained for the moving boundary and temperatures, the effects of several parameters such as latent heat, Boltzmann constant, density ratios, heat transfer coefficients, etc. have been shown. The correctness of numerical results has also been checked by satisfying the integral heat balance at every timestep.

Formation of Single-Phase Indium Selenide Thin Films by Elemental Evaporation

Formation of crystalline, monophasic indium selenide (InSe) thin solid films by elemental evaporation on hot glass substrates (400 to 530 K) is reported. The compound formation as well as the composition of the formed films are confirmed by X-ray photoelectron spectroscopy (XPS) analysis. The crystallinity of the rhombohedral InSe films can be improved by post-depositional annealing for t < 30 min at 533 K. The InSe thin films become Se-deficient at higher temperatures of deposition or longer duration of annealing. Optical studies reveal the bandgap to be around 1.29 eV. Under optimum conditions of preparations the lowest resistivity of ≈ 12.8 Ω cm is obtained. Durch Verdampfen aus den Elementen auf heiße Glassubstrate (400 bis 530 k) werden dünne, kristalline, einphasige Indiumselenid (InSe)-Festkörperschichten gebildet. Sowohl die Bildung der Verbindung als auch die Zusammensetzung der Schichten werden durch Röntgen-Photoelektronenspektroskopie (XPS) untersucht. Die Kristallinität der rhomboedrischen InSe-Schichten kann durch eine Temperung bei 533 K für t < 30 min nach der Abscheidung verbessert werden. Die dünnen InSe-Schichten zeigen nach Abscheidung bei höheren Temperaturen oder längerer Temperungsdauer einen Se-Unterschuß. Optische Untersuchungen ergeben, daß die Bandlücke bei etwa 1,29 eV liegt. Unter optimalen Präperationsbedingungen wird ein niedrigster Widerstand von ≈ 12.8 Ω cm erreicht.

Phase Transformation Introduced by Mechanical and Chemical Surface Preparations of Tetragonal Zirconia Polycrystals

Cutting of Y2O3-doped TZP rods by a low-speed diamond saw introduces an unidentified, metastable phase X (x-ZrO2) coexisting with the tetragonal (t-ZrO2) and the monoclinic (m-ZrO2) phases initially present in the sample. Further mechanical deformation of the cut surface by indentation or polishing sustains the x-ZrO2. Chemical etching removes the x-ZrO2 and increases the m-ZrO2content.

CDNA Cloning, Overexpression in Escherichia coli, Purification and Characterization of Sheep Liver Cytosolic Serine Hydroxymethyltransferase

A sheep liver cDNA clone for the cytosolic serine hydroxymethyltransferase (SHMT) was isolated and its nucleotide sequence determined. The full-length cDNA of SHMT was placed under the control of T7 promoter in pET-3C plasmid and expressed in Escherichia coli. The overexpressed enzyme, present predominantly in the soluble fraction, was catalytically active. The recombinant SHMT was purified to homogeneity with a yield of 10 mg/l bacterial culture. The recombinant enzyme was capable of carrying out tetrahydrofolate-dependent and tetrahydrofolate-independent reactions as effectively as the native enzyme. The K-m values for serine (1 mM) and tetrahydrofolate (0.82 mM) were similar to those of the native enzyme. The recombinant enzyme had a characteristic visible spectrum indicative of the presence of pyridoxal 5'-phosphate as an internal aldimine. The apoenzyme obtained upon removal of the cofactor was inactive and could be reconstituted by the addition of pyridoxal 5'-phosphate demonstrating that the recombinant SHMT was functionally very similar to the native SHMT. This overexpression of eukaryotic tetrameric SHMT in E. coli and the purification and characterization of the recombinant enzyme should thus allow studies on the role of specific amino acids and domains in the activity of the enzyme.