Identification and confirmation of traces of chlorinated fatty acids in fish downstream of bleached kraft pulp mills by gas chromatography with halogen-specific detection

Methyl esters of threo-9,10-dichlorooctadecanoic, threo-7,8-dichlorohexadecanoic, and threo-5,6-dichlorotetradecanoic acids, present in transesterified extracts of filets, gonad, intestinal fat and carcass of white sucker (Catostomus commersoni) sampled in receiving waters of bleached kraft pulp mill effluents, were identified by gas chromatography with halogen-specific detection (XSD). Identification was based on (1) a comparison of the retention times of a sample peak with a prospective reference standard on two stationary phases of very different polarities by spiking, and (2) elution behavior of configurational and positional isomers of dichloro fatty acid methyl esters.     

Vibrational spectra and rotational isomerism of simple dialkyl dithioesters and N-acylglycine ethyl dithioesters,thiolesters, and dioxygenesters

Simple dialkyl dithioesters and N-acylglycine ethyl dithioesters have been investigated by Raman, resonance Raman and infrared spectroscopies. Based on the data of isotopic substituted derivatives and a normal coordinate analysis, vibrational assignments have been proposed for methyl dithioacetate. Raman spectra of ethyl dithioacetate(CH₃C(=S)S

CH₂CH₃) and methyl dithiopropionate (CH₃CH₂

C (=S)SCH₃) have provided fundamental knowledge on rotational isomerism about the bonds indicated. Raman, resonance Raman and FTIR spectroscopic studies on N-acylglycine ethyl dithioesters indicated the coexistence of three conformers in solution and a combined x-ray crystallographic-resonance Raman spectroscopic approach has been used to set up precise structure-spectra correlations for these conformers. The corresponding conformational states of N-acylglycine thiolesters and dioxygenesters are also discussed briefly.     

Two-dimensional electrophoresis reveals differential protein expression in high- and low-secreting variants of the rat basophilic leukaemia cell line

The aim of this investigation was the identification of cellular proteins that confer a high secretory phenotype on subclones of the rat basophilic leukaemia (RBL) cell line as a model of mast cell regulated degranulation. Following protein separation by two-dimensional (2-D) electrophoresis and silver staining, more than 2000 polypeptide "spots" were resolved reproducibly. Higher sample loads and Coomassie blue staining facilitated the identification by delayed extraction-matrix-assisted laser desorption/ionization (DE-MALDI) mass spectrometry of several polypeptides that were differentially expressed in the high- and low-secreting clones. Several proteins were identified whose expression could contribute to the difference in secretory phenotype. Furthermore, silver-stained 2-D gel patterns suggested differential expression of proteins in the 20-25 kDa and the pI 4.5-7.5 range, characteristic of small guanosine 5'-triphosphate (GTP)-binding proteins. By a combination of "GTP overlay" and immunoblotting, we were able to demonstrate differential expression of small GTP binding-proteins, including Rab3 proteins, in high-and low-secreting clones. The sensitivity of this complementary approach facilitated the detection of some GTP binding and Rab3 proteins, whose expression was not evident in silver-stained 2-D gels.     

Blue ferrocenium azurin: an organometalloprotein with tunable redox properties

A ferrocene derivative (2-[(methylsulfonyl)thio]ethylferrocene) (1) has been synthesized and incorporated into apo-azurin from Pseudomonas aeruginosa by covalent attachment to the highly conserved Cys112. The resulting artificial organometalloprotein (a protein containing organometallic compounds in the active site) has been characterized by UV-vis, electrospray mass spectrometry, and cyclic voltammetry (CV). Incorporation of 1 into azurin resulted in a higher solubility of the ferrocene group and improved stability of the ferrocenium species in aqueous solution, as shown by a more intense UV-vis absorption and a more reversible CV of the attached ferrocene group, respectively. The incorporation of 1 also increased the reduction potential of the complex from 402 to 579 mV (vs NHE), consistent with the ferrocene group being encapsulated inside the hydrophobic environment of the protein. Modulation of the reduction potential of ferrocene by residues near the secondary coordination sphere has also been demonstrated. Raising the pH from 4 to 9 resulted in a greater than 80 mV decrease in reduction potential of the protein-bound ferrocene (from 579 to 495 mV), while replacing Met121, an amino acid residue in close proximity to the ferrocene group with a positively charged Arg or negatively charged Glu, resulted in the predicted increase or decrease in reduction potential at all pH values. Similarly, substitution of Met121 with a more hydrophobic Leu raised the reduction potential. The increased solubility, stability, and tune-ability of this organometalloprotein make it an ideal choice for carrying out a number of biological reactions, such as long-range electron transfer or sensing. As an example of such applications, stoichiometric oxidation of ferrocytochrome c by the blue ferrocenium azurin was demonstrated.