Characterization of protein contributions to cobalt-carbon bond cleavage catalysis in adenosylcobalamin-dependent ethanolamine ammonia-lyase by using photolysis in the ternary complex

Protein contributions to the substrate-triggered cleavage of the cobalt-carbon (Co-C) bond and formation of the cob(II)alamin-5'-deoxyadenosyl radical pair in the adenosylcobalamin (AdoCbl)-dependent ethanolamine ammonia-lyase (EAL) from Salmonella typhimurium have been studied by using pulsed-laser photolysis of AdoCbl in the EAL-AdoCbl-substrate ternary complex, and time-resolved probing of the photoproduct dynamics by using ultraviolet-visible absorption spectroscopy on the 10(-7)-10(-1) s time scale. Experiments were performed in a fluid dimethylsulfoxide/water cryosolvent system at 240 K, under conditions of kinetic competence for thermal cleavage of the Co-C bond in the ternary complex. The static ultraviolet-visible absorption spectra of holo-EAL and ternary complex are comparable, indicating that the binding of substrate does not labilize the cofactor cobalt-carbon (Co-C) bond by significantly distorting the equilibrium AdoCbl structure. Photolysis of AdoCbl in EAL at 240 K leads to cob(II)alamin-5'-deoxyadenosyl radical pair quantum yields of <0.01 at 10(-6) s in both holo-EAL and ternary complex. Three photoproduct states are populated following a saturating laser pulse, and labeled, P(f), P(s), and P(c). The relative amplitudes and first-order recombination rate constants of P(f) (0.4-0.6; 40-50 s(-1)), P(s) (0.3-0.4; 4 s(-1)), and P(c) (0.1-0.2; 0) are comparable in holo-EAL and in the ternary complex. Time-resolved, full-spectrum electron paramagnetic resonance (EPR) spectroscopy shows that visible irradiation alters neither the kinetics of thermal cob(II)alamin-substrate radical pair formation, nor the equilibrium between ternary complex and cob(II)alamin-substrate radical pair, at 246 K. The results indicate that substrate binding to holo-EAL does not "switch" the protein to a new structural state, which promptly stabilizes the cob(II)alamin-5'-deoxyadenosyl radical pair photoproduct, either through an increased barrier to recombination, a decreased barrier to further radical pair separation, or lowering of the radical pair state free energy, or a combination of these effects. Therefore, we conclude that such a change in protein structure, which is independent of changes in the AdoCbl structure, and specifically the Co-C bond length, is not a basis of Co-C bond cleavage catalysis. The results suggest that, following the substrate trigger, the protein interacts with the cofactor to contiguously guide the cleavage of the Co-C bond, at every step along the cleavage coordinate, starting from the equilibrium configuration of the ternary complex. The cleavage is thus represented by a diagonal trajectory across a free energy surface, that is defined by chemical (Co-C separation) and protein configuration coordinates.     

Growth of aligned carbon nanofibres over large areas using colloidal catalysts at low temperatures

Highly purified cobalt colloids have been employed as a catalyst to grow aligned carbon nanofibres at temperatures as low as 300 degree C by dc plasma enhanced chemical vapour deposition over large areas.     

Circular logic: nonribosomal peptide-like macrocyclization with a ribosomal peptide catalyst

A protease from ribosomal peptide biosynthesis macrocyclizes diverse substrates, including those resembling nonribosomal peptide and hybrid polyketide-peptide products. The proposed mechanism is analogous to thioesterase-catalyzed chemistry, but the substrates are amide bonds rather than thioesters.     

Aspects of stereocontrol in the L-Selectride reduction of 4-acyl-1,3-dioxolane derivatives

The application of L-Selectride, either alone or in combination with ZnCl₂, to aryl ketones 1, 8 and 11 resulted in highly anti-stereoselective reduction. In contrast, lactols 22 and 23 gave a moderate syn-preference using L-Selectride alone and a high syn-preference in the presence of ZnCl₂. Uniquely, high anti- stereoselectivity was observed in the reduction of o-anisyl lactol 37 with L-Selectride alone, which was switched to a high syn-preference when ZnCl₂ was present.     

Inactivation of Virulent Burkholderia pseudomallei by Sunlight

The goal of this study was to determine the sensitivity of virulent Burkholderia pseudomallei to natural sunlight. We describe solar dosimetry calibrated to integrate radiation between 295 and 305 nm and an exposure system that minimizes thermal effects on bacterial cells. Burkholderia pseudomallei cells were either exposed to sunlight in UV transparent dishes or maintained in the dark covered by opaque foil. The cells maintained in the dark remained at constant levels for the duration of all experiments. The exposed cells nearby were killed with a kinetic studied through 5 Log₁₀ inactivation. We found that cells in stationary phase of growth were nearly two-fold more resistant to sunlight than cells in lag or exponential growth. A virulent strain of B. pseudomallei that produced mucoid colonies showed sensitivity to sunlight similar to both a virulent strain that produced nonmucoid colonies and a strain of B. thailandensis . The inactivation of B. pseudomallei by sunlight in different types of water of environmental relevance or inside amoebae was investigated. The sensitivity of virulent B. pseudomallei was calculated and its comparison with previous studies employing monochromatic germicidal light (254 nm) is discussed . This may be the first report in the open literature of the inactivation of a virulent biological threat agent by natural sunlight. These data should assist in estimating the risk for contracting melioidosis and in predicting the time period during which B. pseudomallei remains infectious after an accidental or intentional release in the environment.     

Analysis of the toenail as a biomonitor of supranutritional intake of Zn,Cu, and Mg

The toenail was examined as a biological monitor of Mg, Zn, and Cu intake using an observational case control model. The One Source Cohort matched 63 individuals in Columbia Missouri who took the One Source multivitamin with 63 control individuals. The matching criteria were based on age, sex, ethnicity, smoking status, and body mass index. The multivitamin contained supra-nutritional levels of Se, Mg, Cu, and Zn. The toenail clippings were examined for these elements using instrumental neutron activation analysis (INAA). A statistical analysis did not indicate a significant difference for Mg, Cu, or Zn between the nails of One Source supplement users and control subjects (p<0.76, 0.55, and 0.85, respectively). The trace nutrient Se was used as an internal control. Previous studies have consistently demonstrated that toenail Se is positively correlated with Se supplement use and the analysis did result in a significant correlation in the toenails of One Source users and control subjects (p<1·10⁻⁴). This internal Se control suggests that the One Source Cohort is largely free from misclassification errors that could interfere with the biomonitor response for the supranutritional intake of Mg, Cu, and Zn.     

Electronic spectra of gas-phase polycyclic aromatic nitrogen heterocycle cations: isoquinoline+ and quinoline+

Electronic spectra of the gas-phase isoquinoline(+)-Ar and quinoline(+)-Ar complexes are recorded using photodissociation spectroscopy by monitoring the Ar loss channel. The D(3)←D(0) and D(4)←D(0) band origins for isoquinoline(+)-Ar are observed at 15245 ± 15 cm(-1) and 21960 ± 15 cm(-1), respectively, whereas for quinoline(+)-Ar they appear at 16050 ± 15 cm(-1) and 21955 ± 15 cm(-1), respectively. Strong vibronic progressions for the D(3)←D(0) band systems of both isoquinoline(+)-Ar and quinoline(+)-Ar are modeled and assigned in terms of ring deformation and carbon-carbon stretch vibrational modes using time-dependent density functional theory calculations in conjunction with Franck-Condon simulations. The properties of the isoquinoline(+) and quinoline(+) molecules are compared with those of the isoelectronic naphthalene(+) molecule. The existence of strong progressions in the visible spectra of isoquinoline(+)-Ar and quinoline(+)-Ar suggests that the corresponding isoquinoline(+) and quinoline(+) molecular cations are unlikely to be responsible for diffuse interstellar bands.     

Proteomic analysis of the Arabidopsis thaliana cell wall

With the completion of the Arabidopsis genome, many hypothetical proteins have been predicted without any information on their expression, subcellular localisation and function. We have performed proteomic analysis of proteins sequentially extracted from enriched Arabidopsis cell wall fractions and separated by two-dimensional gel electrophoresis (2-DE). The proteins were identified by peptide mass fingerprinting using matrix-assisted laser desorption/ionisation-time of flight (MALDI-TOF) mass spectrometry and genomic database searches. This is part of a targeted exercise to establish the entire Arabidopsis secretome database. We report evidence for new proteins of unknown function whose existence had been predicted from genomic sequences and, furthermore, localise them to the cell wall. In addition, we observed an unexpected presence in the cell wall preparations of proteins whose known biochemical activity has never been associated with this compartment hitherto. We discuss the implications of these findings and present results suggesting a possible involvement of cell wall kinases in plant responses to pathogen attack.     

The origin of electrochemical activity in Li2MnO3

The electrochemical activity of Li2MnO3 in non-aqueous media has been investigated and found to involve neither Mn(4+)-Mn5+ oxidation nor simultaneous O2- removal but exchange of Li+ by H+, the latter being generated in the electrolyte.     

Caminoside A,an antimicrobial glycolipid isolated from the marine sponge Caminus sphaeroconia

Extracts of the marine sponge Caminus sphaeroconia showed potent activity in a screen for bacterial type III secretion inhibitors. Bioassay guided fractionation of the extract led to the isolation of the novel antimicrobial glycolipid caminoside A (1). The structure of caminoside A was elucidated by analysis of spectroscopic data and chemical degradation.[structure: see text]