Cloning, sequencing, and expression of the meso-diaminopimelate dehydrogenase gene from Bacillus sphaericus

The gene encoding the meso-diaminopimelate dehydrogenase of Bacillus sphaericus was cloned into E. coli cells and its complete DNA sequence was determined. The meso-diaminopimelate dehydrogenase gene consisted of 978 nucleotides and encoded 326 amino acid residues corresponding to the subunit of the dimeric enzyme. The amino acid sequence deduced from the nucleotide sequence of the enzyme gene of B. sphaericus showed 50% identity with those of the enzymes from Corynebacterium glutamicum and Brevibacterium flavum. The enzyme gene from B. sphaericus was highly expressed in E. coli cells. We purified the enzyme to homogeneity from a transformant with 76% recovery. The N-terminal amino acid of both the enzyme from B. sphaericus and the transformant were serine, indicating that the N-terminal methionine is removed by post-translational modification in B. sphaericus and E. coli cells.     

Catalytic chemistry of solid polyoxometalates and their industrial applications

First, fundamental properties (structure, acid and redox properties) and advantages of solid polyoxometalate catalysts (catalyst design by acid and redox control, molecularity, unusual reaction field and unique basicity) are explained. Then, the mechanism of alcohol dehydration elucidated by direct observation of reaction intermediates by solid-state NMR and the very high activity of Cs_2.5H_0.5PW₁₂O₄₀ are described. Finally several industrial applications of polyoxometalate catalysts are briefly introduced placing stress on the role of unique chemical properties of polyoxometalates.     

New signal transduction mechanisms of atrial natriuretic factor: inhibition of phosphorylation of protein kinase C and A 240 kDa protein in adrenal cortical plasma membrane by cGMP dependent and independent mechanisms

The effects of synthetic atrial natriuretic factor (ANF) on the state of protein phosphorylation in plasma membranes of bovine adrenal cortex have been studied in vitro. ANF (1x10(-8)M - 1x10(-7)M) specifically inhibited the phosphorylation of two distinct proteins of 78 kDa and 240 kDa. Immunoblotting with specific antiserum to protein kinase C produced evidence that 78 kDa protein is most likely the protein kinase C whose phosphorylation is inhibited by both ANF and cGMP. However, cGMP did not affect the phosphorylation of 240 kDa protein, indicating a new cGMP-independent mechanism of ANF action in the adrenal, which is compatible with the lack of action of cGMP and its analogs in ANF-induced inhibition of aldosterone secretion from adrenal cortex. The inhibition of phosphorylation of putative protein kinase C by ANF or cGMP indicates a hitherto unknown signal transduction mechanism of ANF.     

Evidence for extracellular localization of activator calcium in dog coronary artery smooth muscle as studied by the pyroantimonate method

Summary Correlated physiological and electron-microscopic studies were made on the source of calcium activating the contractile system (activator calcium) in dog coronary artery smooth muscle fibers. The magnitude of contracture tension induced by 100 mM K⁺ was dependent on external Ca²⁺ concentration and reduced or eliminated by factors known to reduce the Ca²⁺ spike or ca²⁺ influx. Little or no mechanical response was elicited by treatments known to cause release of intracellularly stored calcium. These results indicated that the contractile system is mainly activated by the inward movement of extracellular calcium. In accordance with the physiological experiments, electron-opaque pyroantimonate precipitate containing calcium was found in the lumina of caveolae, but not in any intracellular structures close to the plasma membrane, when the relaxed fibers were fixed in a 1% osmium tetroxide solution containing 2% potassium pyroantimonate. If the contracted fibers were fixed in the same solution, the pyroantimonate precipitate was diffusely distributed in the myoplasm in the form of numerous particles, while the precipitate in the caveolar lumina was scarcely seen. These findings are discussed in connection with the regulation of intracellular Ca²⁺ concentration in dog coronary artery smooth muscle.     

The Assignment of the 13C-NMR Spectrum of Lysocellin and Its Biosynthesis

The complete ¹⁸C-NMR assignment of lysocellin sodium salt has been obtained based on the ¹³C-¹³C double labeling method and comparison with its retroaldol degradation product as well as a structurally related polyether antibiotic, lasalocid A.

In parallel, the biosynthesis of lysocellin was investigated by feeding ¹³C-labeled precursors followed by analyzing the resulting labeling patterns of the ¹³C-NMR spectra; a pathway to the antibiotic molecule is suggested which proceeds through condensation of two butyrate, eight propionate and one acetate units.

In addition, a conversion of butyrate to propionate during the metabolic process has been disclosed.     

Virulence phenotypes of Shigella flexneri 2a avirulent mutant 24570 can be complemented by the plasmid-coded positive regulator virF gene

Avirulent mutation of an opaque colony variant of Shigella flexneri 2a designated 24570 has been believed to be linked with the glpK locus of the chromosome. However, avirulent phenotypes of the 24570 strain could be complemented by the invasion plasmid-coded virF gene, a positive regulator for invasion genes. The 24570 strain had a DNA structural alteration upstream of the virF gene.