Characterization of HgCl2 Tridentate Amine Complexes by X-ray Crystallography, NMR and ESI-MS

Two new HgCl2 complexes of tridentate nitrogen ligands were characterized by X-ray crystallography, proton NMR spectroscopy and ESI-MS. The five-coordinate complex [Hg(BMPA)Cl-2] (1) (BMPA = bis(2-pyridylmethyl)amine) crystallized from acetonitrile/m-xylene by slow evaporation in the monoclinic space group P2(1)/n with a = 8.3896(8) , b = 12.8020(13) , c = 13.3526(13) , alpha = 90A degrees, beta A = 90.480(2)A degrees, gamma A = 90A degrees and z = 4. The square pyramidal structure (tau = 0.009) has approximate C (s) symmetry. Despite comparable Hg-N bond lengths in 1, inversion of the central nitrogen was rapid on the chemical shift time scale in dilute solution except at very low temperatures. The related complex [Hg(BEPA)Cl-2] (2) (BEPA = bis(2-{pyrid-2-yl}ethyl)amine) crystallized from acetonitrile/ethyl acetate/hexanes by slow diffusion in the orthorhombic space group Pnma with a = 13.424(3) , b = 14.854(3) , c = 8.118(2) , alpha = 90A degrees, beta A = 90A degrees, gamma A = 90A degrees and z = 4. The mixed geometry structure (tau = 0.56) also has crystallographic mirror symmetry as well as C (s) point group symmetry. In dilute acetonitrile solution, 1 was stable while 2 slowly converted to a more thermodynamically stable complex.

Characterization of vascular cavity coatings and microscopic tube-shaped structures from Upper Cretaceous dinosaur bone: evidence of a bacterial origin for dinosaur blood vessels""

Recent claims of blood vessels extracted from dinosaur fossils challenge classical views of soft-tissue preservation. Alternatively, these structures may represent postdepositional,diagenetic biofilms that grew on vascular cavity surfaces within the fossil. Similar red, hollow, tube-shaped structures were recovered from well-preserved and poorly-preserved (abraded, desiccated, exposed) Upper Cretaceous dinosaur fossils in this study. Integration of light microscopy, scanning electron microscopy, and energy dispersive x-ray spectroscopy was used to compare these vessel structures to the fossils from which they are derived. Vessel structures are typically 100-400 μm long, 0.5-1.5 μm thick, 10-40 μm in diameter and take on a wide range of straight, curved, andbranching morphologies. Interior surfaces vary from smooth to globular and typically contain spheres, rods, and fibrous structures (< 2 μm in diameter) incorporated into the surface. Exterior surfaces exhibit 2-μm-tall converging ridges, spaced 1-3 μm apart, that are sub-parallel to the long axis of the vessel structure. Fossil vascular cavities are typically coated with a smooth or grainy orange layer that shows a wide range of textures including smooth, globular, rough, ropy, and combinations thereof. Coatings tend to overlay secondary mineral crystals and framboids, confirming they are not primary structures of the fossil. For some cavity coatings, the surface that had been in contact with the bone exhibits a ridged texture, similar to that of vessel structures, having formed as a mold of the intravascular bone surface. Thus, vessel structures are interpreted as intact cavity coatings isolated after the fossil is demineralized. The presence of framboids and structures consistent in size and shape with bacteria cells, the abundance of iron in cavity coatings, and the growth of biofilms directly from the fossil that resemble respective cavity coatings support the hypothesis that vessel structures result from ironconsuming bacteria that form biofilms on the intravascular bone surfaces of fossil dinosaur bone. This also accounts for microstructures resembling osteocytes as some fossil lacunae are filled with the same iron oxide that comprises vessel structures andcoatings. Results of this study show that systematic, high-resolution SEM analyses of vertebrate fossils can provide improved insight on microtaphonomic processes, including the role of bacteria in diagenesis. These results conflict with earlier claims of dinosaurblood vessels and osteocytes.

Hydrodynamic Characterization of Bile Salt Host-Guest Complexes by Pulsed Field Gradient NMR Spectroscopy

The work described herein is aimed at understanding primary and secondary aggregation of bile salt micelles and how micelles can perform chiral recognition of binapthyl analytes. Previous work with cholate and deoxycholate using micellar electrokinetic chromatography (MEKC) and nuclear magnetic resonance (NMR) has provided insightinto cholate and deoxycholate micelle formation, especially with respect to the critical micelle concentration (CMC). Chiral separations of the model analyte, 1,1â??-binaphthyl-2,2â??-diyl hydrogen phosphate (BNDHP), via cholate (C) and deoxycholate (DC) mediated MEKC separataions previously have shown the DC CMC to be 7-10 mM andthe cholate CMC at 14 mM at ph 12. A second model analyte,1,1â??-binaphthol (BN), was also previously investigated to probe micellar structure, but the MEKC data for this analyte implied a higher CMC, which may be interpreted as secondary aggregation. Thiswork extends the investigation of bile salts to include pulsed field gradient spin echo (PFGSE) NMR experiments being used to gain information about the size and degree of polydispersity of cholate and deoxycholate micelles. Concentrations of cholate below 10mM show a large variation in effective radius likely due to the existence of transient preliminary aggregates. The onset of the primary micelle shows a dramatic increase in effective radius of the micelle in cholate and deoxycholate. In the region of expectedsecondary aggregation a gradual increase of effective radius was observed with cholate; deoxycholate showed a persistent aggregate size in the secondary micelle region that is modulated by the presence of an analyte molecule. Effective radii of cholate anddeoxycholate (individually) were compared with and without R- and S-BNDHP in order to observe the effective radius difference of micelles with and without analyte present. The presence of S-BNDHP consistently resulted in a larger effective aggregate radius incholate and deoxycholate, confirming previous data of the S-BNDHP interacting more with the micelle than R-BNDHP. In total, various NMR techniques, like diffusion NMR can be used to gain a greater understanding of the bile salt micellization process and chiral resolution.

Purification and Preliminary Characterization of the TM0727 Protein from Thermotoga maritima

The TM0727 gene of Thermotoga maritima is responsible for encoding what has been reported to be a modulator of DNA gyrase (pmbA). Although the function of pmbA is still unknown, it is believedto be involved in cell division, carbon storage regulation, and the synthesis of the antibiotic peptide microcin B17. It is suggested that it serves together with tldD, a known zinc dependent protease, tomodulate DNA gyrase. TM0727 is believed to be a zinc dependent protease that binds zinc in the central active site of the molecule, located between two equivalent monomeric units. However, thecrystal structure determined by Wilson et al. (2005) did not contain zinc. It therefore remains to be seen if TM0727 requires zinc for activity, or regulation, and if the protein is indeed a protease. To begin studying this protein, the gene was expressed in BL21(DE3) pLysS cells and the induction time was optimized. Using affinity and ion exchange chromatography, the protein has been successfully purified. The purification procedure can be replicated to obtain sufficient protein for characterization. Purification results show that the protein loses stability after 24 hours and remains stable under an imidazole-free lysis workup. Preliminary characterization of TM0727 has focused on understanding the protein’s structuralproperties through tryptophan fluorescence anisotropy measurements. The four tryptophan residues located within the TM0727 dimer fluoresce at different maximum wavelengths and with differentintensities upon excitation with 295nm light. These emission properties are highly sensitive to the environment (solvent, surrounding residues) of each tryptophan residue. The low number oftryptophans allows for a specific monitoring of the protein’s structure as it denatures. As more denaturant is added to the protein, its tryptophan environments have clearly altered. This is indicative of unfolding and increased solvent exposure of the protein. This unfolding has been confirmed with the addition of a fluorescent quencher. Additionally, fluorescence anisotropy measurements have been carried out on the protein to gain a preliminary understanding of the rotational dynamics of the tryptophan residues. These experiments excite the tryptophan residues within the sample using a polarized light source. Polarized emission is then detected, the degree of which depends on the rotational dynamics and local environment of the tryptophan residues. The protein was denatured and the changes in emission were recorded to detect these structural changes. Results have shown a large change in quaternary structure, consistent with a dimer to monomer transition, occurs at 1.5M Guandidine HCl. There has also been an examination of the crystal structure for the location of a potential active site. The inner cavity of the protein was inspected visually to locate a potential location for a catalytic triad, specifically the amino acids found in the active sites of serine, cyteine, and aspartateproteases. It was found that a potential aspartic protease active site may be located between the Asparate286 and Aspartate287 residues. Further investigation is warranted to test this remotepossibility.