Desulfurization of model and diesel oils by resting cells of Gordona sp.

The desulfurization activity of the resting cells of Gordona sp. CYKS1 was strongly depended on harvest time and the highest value when the cells had been harvested in the early growth phase (0.12 mg sulfur g^–1 cell^–1 h^–1). For the model oil, hexadecane containing dibenzothiophene, the specific desulfurization rate decreased as the reaction proceeded. Both the specific and the volumetric desulfurization rates were not significantly affected by the aqueous-to-oil phase ratio. The diesel oils, light gas oil and a middle distillate unit feed were desulfurized at higher rates (ca. 0.34 mg sulfur g^–1 cell^–1 h^–1) than the model oil (0.12 mg sulfur g^–1 cell^–1 h^–1).     

An Insertion Peptide in Yeast Glycyl-tRNA Synthetase Facilitates both Productive Docking and Catalysis of Cognate tRNAs

The yeast Saccharomyces cerevisiae possesses two distinct glycyl-tRNA synthetase (GlyRS) genes: GRS1 and GRS2. GRS1 is dually functional, encoding both cytoplasmic and mitochondrial activities, while GRS2 is dysfunctional and not required for growth. The protein products of these two genes, GlyRS1 and GlyRS2, are much alike but are distinguished by an insertion peptide of GlyRS1, which is absent from GlyRS2 and other eukaryotic homologues. We show that deletion or mutation of the insertion peptide modestly impaired the enzyme''s catalytic efficiency in vitro (with a 2- to 3-fold increase in K_m and a 5- to 8-fold decrease in k_cat). Consistently, GRS2 can be conveniently converted to a functional gene via codon optimization, and the insertion peptide is dispensable for protein stability and the rescue activity of GRS1 at 30°C in vivo. A phylogenetic analysis further showed that GRS1 and GRS2 are paralogues that arose from a gene duplication event relatively recently, with GRS1 being the predecessor. These results indicate that GlyRS2 is an active enzyme essentially resembling the insertion peptide-deleted form of GlyRS1. Our study suggests that the insertion peptide represents a novel auxiliary domain, which facilitates both productive docking and catalysis of cognate tRNAs.     

A Yersinia pseudotuberculosis protein which cross-reacts with HLA-B27

The most-debated question in the investigation of the spondyloarthropathies has been whether there is molecular mimicry between host HLA-B27 antigens and the arthritis-causing pathogens. We have generated a monoclonal anti-HLA-B27 antibody in our laboratory and have used a radioimmunoassay to screen a panel of bacterial species. Two strains of Yersinia pseudotuberculosis were found to be highly reactive. The cross-reactive Yersinia component was identified by Western blot to be a 19,000 component. A preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis chromatography apparatus was constructed to isolate milligram quantities of this component. To verify that the component carried the HLA-B27-specific epitope, rabbits were hyperimmunized with the purified materials. Affinity-purified antibodies from one of the immunized rabbits indeed carried anti-HLA-B27 activity. Last, antibodies generated against synthetic peptides derived from the HLA-B27.1 amino acid sequence were tested against the Yersinia component. Positive reactivity was found with antibodies generated against a peptide spanning residues 69-83 of the HLA-B27.1 protein. Since this resides in the segment responsible for the allotypic specificity of the antigen, these experiments establish the presence of molecular mimicry to a high degree of confidence.     

Extracellular secretion of levansucrase from Zymomonas mobilis in Escherichia coli

Secretion of levansucrase from Zymomonas mobilis in Escherichiacoli by glycine supplement was investigated. A significant amount of levansucrase (about 25% of total activity) was found in intact whole-cells. Cell fractionation experiments showed that levansucrase was found both in the periplasmic space and in the cytoplasmic fraction of E. coli. None or only trace amounts of levansucrase was detected in the extracellular culture broth at 24 h of cultivation and it accrued with the increasing concentration of glycine in the culture medium and duration of the culture period. Optimal glycine concentration for the maximum secretion of levansucrase was in the range of 0.8-1%, in which approximately 20-50% of levansucrase was released into the extracellular fraction at 24 h of cultivation, although glycine retarded the bacterial growth.     

Towards an integrated system for bio-energy: hydrogen production by <Emphasis Type="Italic">Escherichia coli</Emphasis> and use of palladium-coated waste cells for electricity generation in a fuel cell

Escherichia coli strains MC4100 (parent) and a mutant strain derived from this (IC007) were evaluated for their ability to produce H₂ and organic acids (OAs) via fermentation. Following growth, each strain was coated with Pd(0) via bioreduction of Pd(II). Dried, sintered Pd-biomaterials (‘Bio-Pd’) were tested as anodes in a proton exchange membrane (PEM) fuel cell for their ability to generate electricity from H₂. Both strains produced hydrogen and OAs but ‘palladised’ cells of strain IC007 (Bio-Pd_IC007) produced ~threefold more power as compared to Bio-Pd_MC4100 (56 and 18 mW respectively). The power output used, for comparison, commercial Pd(0) powder and Bio-Pd made from Desulfovibrio desulfuricans, was ~100 mW. The implications of these findings for an integrated energy generating process are discussed.     

Immunocytochemical localization of tubulin with colloidal gold in cilia of rabbit tracheal epithelial cultures

Ciliated tracheal epithelia cell cultures were investigated immunocytochemically with anti-tubulin and colloidal gold. When rabbit tracheal cultures were fixed in paraformaldehyde, treated with acetone, anti-tubulin and a second antibody coupled to FITC, fluorescence was associated with cytoskeletal and axonemal microtubules. Cilia covering the apical surface of the ciliated tracheal cells fluoresced very brightly thus facilitating identification of this cell type. Electron microscopy of tracheal cultures fixed as above, treated with Triton-X 100 and incubated in anti-tubulin and protein A coupled to colloidal gold resulted in the highly specific localization of tubulin in ciliary axonemes and basal bodies. Omission of primary or secondary antibody resulted in extremely low levels of fluorescence while no colloidal gold particles could be detected in cultures at the electron microscopy level when rabbit anti-tubulin was omitted.     

Inhibitory Effect of Glycerin on Vibrio parahaemolyticus and Salmonella

In a study of the effect of glycerin in transport media on Vibrio parahaemolyticus and Salmonella, it was found that a concentration of 30% glycerin was highly inhibitory for V. parahaemolyticus and to a lesser degree for Salmonella. The incorporation of peptone or human feces in media did not reduce the inhibitory effect of glycerin. In media with 15% glycerin, viable counts of V. parahaemolyticus and Salmonella increased after 24 hr of incubation both in the presence and absence of feces. Due to the concurrent increase in the total bacterial count in the media containing feces, no enrichment effect was noted.