Deoxyhypusine synthase is phosphorylated by protein kinase C in vivo as well as in vitro

Deoxyhypusine synthase catalyzes the first step in the posttranslational synthesis of an unusual amino acid, hypusine, in the eukaryotic translation initiation factor 5A (eIF-5A) precursor protein. We earlier observed that yeast recombinant deoxyhypusine synthase was phosphorylated by protein kinase C (PKC) in vitro (Kang and Chung, 1999) and the phosphorylation rate was synergistically increased to a 3.5-fold following treatment with phosphatidylserine (P.Ser)/diacylglycerol (DAG)/ Ca(2+), suggesting a possible involvement of PKC. We have extended study on the phosphorylation of deoxyhypusine synthase in vivo in different cell lines in order to define its role on the regulation of eIF5A in the cell. Deoxyhypusine synthase was found to be phosphorylated by endogenous kinases in CHO, NIH3T3, and chicken embryonic cells. The highest degree of phosphorylation was found in CHO cells. Moreover, phosphorylation of deoxyhypusine synthase in intact CHO cells was revealed and the expression of phosphorylated deoxyhypusine synthase was significantly diminished by diacyl ethylene glycol (DAEG), a PKC inhibitor, and enhanced by phorbol 12-myristate 13-acetate (PMA) or Ca(2+)/DAG. Endogenous PKC in CHO cell and cell lysate was able to phosphorylate deoxyhypusine synthase and this modification is enhanced by PMA or Ca(2+) plus DAG. Close association of PKC with deoxyhypusine synthase in the CHO cells was evident in the immune coprecipitation and was PMA-, and Ca(2+)/phospholipid dependent. These results suggest that phosphorylation of deoxyhypusine synthase was PKC-dependent cellular event and open a path for possible regulation in the interaction with eIF5A precursor for hypusine synthesis.     

Improved computer algorithm for characterizing skewed chromatographic band broadening : II. Results and comparisons

A newly developed method using an exponentially modified Gaussian peak shape model produces results that are more precise and less subject to baseline noise than previous methods for characterizing chromatographic band broadening. The method requires only precisely measurable experimental peak parameters: peak retention time, peak height, peak area, and peak centroid (first moment). Accuracy and precision of the new method were compared with other digital approaches by using computer-synthesized peaks and experimental chromatographic data from many HPLC columns. The proposed method offers a reasonable compromise between accuracy, precision, and convenience. A rapid visual estimate of peak skew can be made by inspecting peak shape and referring to a calibration plot involving peak parameters. Peak variance and skew data from this method are also useful for finding column dispersion corrections in size-exclusion chromatography calibrations.     

Photophysics of aromatic molecules with low-lying pi sigma* states: excitation-energy dependence of fluorescence in jet-cooled aromatic nitriles

Excitation-energy dependence of fluorescence intensity and fluorescence lifetime has been measured for 4-dimethylaminobenzonitrile (DMABN), 4-aminobenzonitrile (ABN), 4-diisopropylaminobenzonitrile (DIABN), and 1-naphthonitrile (NN) in a supersonic free jet. In all cases, the fluorescence yield decreases rather dramatically, whereas the fluorescence lifetime decreases only moderately for S1 (pi pi*, L(b)) excess vibrational energy exceeding about 1000 cm(-1). This is confirmed by comparison of the normalized fluorescence excitation spectrum with the absorption spectrum of the compound in the vapor phase. The result indicates that the strong decrease in the relative fluorescence yield at higher energies is due mostly to a decrease in the radiative decay rate of the emitting state. Comparison of the experimental results with the TDDFT potential energy curves for excited states strongly suggests that the decrease in the radiative decay rate of the aminobenzonitriles at higher energies is due to the crossing of the pi pi* singlet state by the lower-lying pi sigma*(C[triple bond]N) singlet state of very small radiative decay rate. The threshold energy for the fluorescence "break-off" is in good agreement with the computed energy barrier for the pi pi*/pi sigma* crossing. For NN, on the other hand, the observed decrease is in fluorescence yield at higher excitation energies can best be attributed to the crossing of the pi pi* singlet state by the pi sigma* triplet state.     

Ground-state properties and static dipole polarizabilities of the alkali dimers from K2 n to Fr2 n(n=0,+1) from scalar relativistic pseudopotential coupled cluster and density functional studies

The newly adjusted energy-consistent nine-valence-electron pseudopotentials for K to Fr are used to calculate spectroscopic properties for the neutral and positively charged alkali dimers using coupled cluster and density functional theory. For the neutral dimers the static dipole polarizability was calculated. The coupled cluster results are all in excellent agreement with experimental values. The density functionals used can give quite different spectroscopic properties especially for the dipole polarizability, with the Perdew-Wang PW91 functional performing best.     

Binding of methyl orange and its homologs by powdered nylon 612: Peculiar temperature dependence on the binding

The ability of powdered Nylon 612 to bind methyl orange, ethyl orange, propyl orange, and butyl orange was investigated at 5, 15, 25 and 35°C in an aqueous solution. The amount of binding of the dye is much higher with this polyamide than with powdered Nylon 66 reported previously,¹ although the former polymer has fewer amide end groups. The Van't Hoff plots of the first binding constant for the binding of butyl orange and propyl orange by powdered Nylon 612 exhibit a bell-shaped curve, whereas the plots for methyl orange and ethyl orange do not. Maximal binding occurs at approximately 15°C for propyl orange and at about 25°C for butyl orange. This is the first instance where the peculiar temperature dependence of the binding constant has been found in the binding of propyl orange, whose hydrophobicity is less than that of butyl orange. These tendencies can be accounted for in terms of increased hydrophobic of butyl orange. These tendencies can be accounted for in terms of increased hydrophobic domains in powdered Nylon 612 and enhanced hydrophobic contributions in the binding process.     

A massively parallel electrochemical approach to the miniaturization of organic micro- and nanostructures on surfaces

This paper describes a simple and convenient strategy for reducing the dimensions of organic micro-and nanostructures on metal surfaces. By varying electrochemical desorption conditions, features patterned by dip-pen nanolithography or micro contact printing and made of linear alkanethiols or selenols can be gradually desorbed in a controlled fashion. The process is referred to as electrochemical whittling because the adsorbate desorption is initiated at the exterior of the feature and moves inward as a function of time. The whittling process and adsorbate desorption were studied as a function of substrate morphology, adsorbate head and tail groups, and electrolyte solvent and salt. Importantly, one can independently address different nanostructures made of different adsorbates and effect their miniaturization based upon ajudicious selection of adsorbate, applied potential, and supporting electrolyte. Some of the physical and chemical origins of these observations have been elucidated.     

Inhibitory effects of 1,3-selenazol-4-one derivatives on mushroom tyrosinase

This study reports depigmenting potency of 1,3-selenazol-4-one derivatives, which would be based upon the finding of direct inhibition to mushroom tyrosinase. 1,3-Selenazol-4-one derivatives exhibited inhibitory effect on dopa oxidase activity of mushroom tyrosinase. In this study, inhibitory effects of six kinds of 1,3-selenazol-4-one derivatives (A, B, C, D, E and F) on mushroom tyrosinase were investigated. Compounds at a concentration of 500 microM exhibited 33.4-62.1% of inhibition on dopa oxidase activity of mushroom tyrosinase. Their inhibitory effects were higher than that of kojic acid (31.7%), a well known tyrosinase inhibitor. 2-(4-Methylphenyl)-1,3-selenazol-4-one (A) exhibited the strongest inhibitory effect among them dose-dependently and in competitive inhibition manner.     

Ortho alkylation of aromatic ketimine with functionalized alkene by Rh(I) catalyst

[reaction: see text] The reaction of the imine of aromatic ketones with functionalized alkenes was performed under a catalytic amount of (PPh3)3RhCl, and corresponding ortho-alkylated ketones were obtained after hydrolysis. A variety of functional groups in the alkene were tolerated in this ortho alkylation. This procedure expands the scope of ortho alkylation to the direct ortho functionalization of aromatic ketones.     

X-ray spectroscopy of enzyme active site analogues and related molecules: bis(dithiolene)molybdenum(IV) and -tungsten(IV,VI) complexes with variant terminal ligands

The X-ray absorption spectra at the molybdenum and selenium K-edges and the tungsten L2,3-edges are acquired for a set of 14 Mo(IV) and W(IV,VI) bis(dithiolene) complexes related to the active sites of molybdo- and tungstoenzymes. The set includes square pyramidal [MoIVL(S2C2Me2)2]- (L = O2-, R3SiO-, RO-, RS-, RSe-) and [WIV(OR)(S2C2Me2)2]-, distorted trigonal prismatic [MoIV(CO)(SeR)(S2C2Me2)2]- and [WIV(CO)L(S2C2Me2)2]- (L = RS-, RSe-), and distorted octahedral [WVIO(OR)(S2C2Me2)2]-. The dithiolene simulates the pterin-dithiolene cofactor ligand, and L represents a protein ligand. Bond lengths are determined by EXAFS analysis using the GNXAS protocol. Normalized edge spectra, non-phase-shift-corrected Fourier transforms, and EXAFS data and fits are presented. Bond lengths determined by EXAFS and X-ray crystallography agree to < or = 0.02 A as do the M-Se distances determined by both metal and selenium EXAFS. The complexes [MoIV(QR)(S2C2Me2)2]- simulate protein ligation by the DMSO reductase family of enzymes, including DMSO reductase itself (Q = O), dissimilatory nitrate reductase (Q = S), and formate dehydrogenase (Q = Se). Edge shifts of these complexes correlate with the ligand electronegativities. Terminal ligand binding is clearly distinguished in the presence of four Mo-S(dithiolene) interactions. Similarly, five-coordinate [ML(S2C2Me2)2]- and six-coordinate [M(CO)L(S2C2Me2)2]- are distinguishable by edge and EXAFS spectra. This study expands a previous XAS investigation of bis(dithiolene)metal(IV,V,VI) complexes (Musgrave, K. B.; Donahue, J. P.; Lorber, C.; Holm, R. H.; Hedman, B.; Hodgson, K. O. J. Am. Chem. Soc. 1999, 121, 10297) by including a larger inventory of molecules with variant physiologically relevant terminal ligation. The previous and present XAS results should prove useful in characterizing and refining metric features and structures of enzyme sites.     

Synthesis and characterization of highly ordered Ni-MCM-41 mesoporous molecular sieves

Highly ordered Ni-MCM-41 samples with nearly atomically dispersed nickel ions were prepared reproducibly and characterized. Similar to the Co-MCM-41 samples, the pore diameter and porosity can be precisely controlled by changing the synthesis surfactant chain length. Nickel was incorporated by isomorphous substitution of silicon in the MCM-41 silica framework, which makes the Ni-MCM-41 a physically stable catalyst in harsh reaction conditions such as CO disproportionation to single wall carbon nanotubes or CO2 methanation. X-ray absorption spectroscopy results indicate that the overall local environment of nickel in Ni-MCM-41 was a tetrahedral or distorted tetrahedral coordination with surrounding oxygen anions. Hydrogen TPR revealed that our Ni-MCM-41 samples have high stability against reduction; however, compared to Co-MCM-41, the Ni-MCM-41 has a lower reduction temperature, and both the H2-TPR and in situ XANES TPR reveal that the reducibility of nickel is not clearly correlated with the pore radius of curvature, as in the case of Co-MCM-41. This is probably a result of nickel being thermodynamically more easily reduced than cobalt. The stability of the structural order of Ni-MCM-41 has been investigated under SWNT synthesis and CO2 methanation reaction conditions as both require catalyst exposure to reducing environments leading to formation of metallic Ni clusters. Nitrogen physisorption and XRD results show that structural order was maintained under both SWNT synthesis and CO2 methanation reaction conditions. EXAFS results demonstrate that the nickel particle size can be controlled by different prereduction temperatures but not by the pore radius of curvature as in the case of Co-MCM-41.