Modeling and analysis of soybean (Glycine max. L) Cu/Zn,Mn and Fe superoxide dismutases

Superoxide dismutase (SOD, EC 1.15.1.1) is an important metal-containing antioxidant enzyme that provides the first line of defense against toxic superoxide radicals by catalyzing their dismutation to oxygen and hydrogen peroxide. SOD is classified into four metalloprotein isoforms, namely, Cu/Zn SOD, Mn SOD, Ni SOD and Fe SOD. The structural models of soybean SOD isoforms have not yet been solved. In this study, we describe structural models for soybean Cu/Zn SOD, Mn SOD and Fe SOD and provide insights into the molecular function of this metal-binding enzyme in improving tolerance to oxidative stress in plants.     

Biotransformation of linear alkylbenzene sulfonate (LAS) by Phanerochaete chrysosporium: oxidation of alkyl side-chain

The white rot fungus Phanerochaete chrysosporium, which generally mineralizes substituted aromatics to CO₂, transformed linear alkylbenzene sulfonate (LAS) surfactants mainly at their alkyl side chain. Degradation of LAS was evidenced by a zone of clearing on LAS-containing agar plates and colorimetric analysis of liquid cultures. Disappearance of LAS was virtually complete within 10 days in low nitrogen (2.4 mM N), high nitrogen (24 mM N) and malt extract (ME) liquid media. After 5 days of incubation in ME medium, transformation of LAS was complete at concentrations4 mg l^-1, but decreased at higher concentrations. The LAS degradation was not dependent on lignin peroxidases (LiPs) and manganese-dependent peroxidases (MnPs). Mineralization of¹⁴C-ring-LAS to ¹⁴CO₂ by P. chrysosporium was <1% regardless of the culture conditions used. Thin layer chromatography and mass spectral analyses indicated that P. chrysosporium transformed LAS to sulfophenyl carboxylates (SPCs) through oxidative shortening of the alkyl side-chains. While LAS disappearance in the cultures was not dependent on LiPs and MnPs, transformation of the parent LAS moieties to SPCs was more extensive in low N medium that favors expression of these enzymes. The SPCs produced in LN cultures were shorter in chain-length than those produced in ME cultures. Also there was a notable shift in the relative abundance of odd and even chain length metabolites compared to the starting LAS particularly in the low N cultures suggesting the possible involvement of processes other than or in addition to-oxidation in the chain-shortening process.     

Genomic fingerprints in the tribe Triticeae produced by PCR using a tRNA consensus primer

Forty-one accessions belonging to ten genera of the tribe Triticeae representing both wild and cultivated species were analyzed by polymerase chain reaction (PCR). Of two consensus tRNA primers tested, one primer revealed characteristic amplification products for all of the species. A total of 35 tRNA markers were scored across all accessions. Five genus-specific and three species-specific markers were obtained. Genomic fingerprints were largely conserved within a genus. The phylogram obtained using parsimony has separated most of the accessions into their prevailing taxonomic species and genus groups. The phylogram showed close association among the three genera Secale, Triticum and Hordeum as expected. The Triticum-Secale relationship was closer than the Triticum-Hordeum or the Secale-Hordeum relationships. The tree also reflected the close associations among the forage grass species belonging to Leymus and Elymus. Thus tDNA-PCR helped to identify species and genera. This revised version was published online in July 2006 with corrections to the Cover Date.     

An in vitro demonstration of peroxisome proliferation and increase in peroxisomal β-oxidation system mRNAs in cultured rat hepatocytes treated with ciprofibrate

Using the normal adult rat hepatocytes, plated on rat tail collagen-coated dishes and fed a chemically defined medium, we demonstrate here that ciprofibrate at 0.1 mM concentration, increases significantly the mRNA levels of fatty acyl-CoA oxidase, enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase bifunctional protein, and thiolase (the three enzymes of the β-oxidation system), and causes peroxisome proliferation. Increase in mRNA levels of these genes was evident within 1 h and was maximal 24 h after the addition of ciprofibrate. In hepatocytes cultured in the absence of ciprofibrate, the basal levels of these enzymes were low and further declined with time. Concomitant treatment of hepatocytes with cycloheximide did not inhibit or superinduce the mRNA levels, indicating that this induction may represent a primary (direct) effect of this compound on the expression of these genes and does not apparently involve short-lived repressor protein(s).     

Fosfomycin Permeation through the Outer Membrane Porin OmpF

Fosfomycin is a frequently prescribed drug in the treatment of acute urinary tract infections. It enters the bacterial cytoplasm and inhibits the biosynthesis of peptidoglycans by targeting the MurA enzyme. Despite extensive pharmacological studies and clinical use, the permeability of fosfomycin across the bacterial outer membrane is largely unexplored. Here, we investigate the fosfomycin permeability across the outer membrane of Gram-negative bacteria by electrophysiology experiments as well as by all-atom molecular dynamics simulations including free-energy and applied-field techniques. Notably, in an electrophysiological zero-current assay as well as in the molecular simulations, we found that fosfomycin can rapidly permeate the abundant Escherichia coli porin OmpF. Furthermore, two triple mutants in the constriction region of the porin have been investigated. The permeation rates through these mutants are slightly lower than that of the wild type but fosfomycin can still permeate. Altogether, this work unravels molecular details of fosfomycin permeation through the outer membrane porin OmpF of E. coli and moreover provides hints for understanding the translocation of phosphonic acid antibiotics through other outer membrane pores.     

Synthesis of novel steroidal D-ring substituted isoxazoline derivatives of 17-oxoandrostanes

A facile synthesis of isoxazoline derivatives of 17-oxoandrostane at the side chain of D-ring is reported. The scheme involves the transformation of the starting dehydroepiandrosterone acetate (ketone) to the Knoevenegel product, reduction to the nitrile, and elimination to the carboxaldehyde. Cycloaddition of nitrileoxides across olefinic aldehyde intermediate led to the synthesis of novel side chain isoxazoline derivatives.     

17-epiestriol,an estrogen metabolite,is more potent than estradiol in inhibiting vascular cell adhesion molecule 1 (VCAM-1) mRNA expression

17-beta estradiol (17-beta E(2)) attenuates the expression of vascular cell adhesion molecule 1 (VCAM-1) in vivo at physiological levels (pg/ml), whereas supraphysiological concentrations of 17-beta E(2) (ng/ml) are required in vitro. We assessed whether a metabolite of estrogen, which could only be generated in vivo, might be a more potent inhibitor of VCAM-1 expression and thereby explain this discrepancy. We report here that 17-epiestriol, an estrogen metabolite and a selective estrogen receptor (ER) beta agonist, is approximately 400x more potent than 17-beta E(2) in suppressing tumor necrosis factor (TNF) alpha-induced VCAM-1 mRNA as well as protein expression in human umbilical vein endothelial cells. Genistein, an ERbeta agonist, at low concentrations (1 and 10 nm) also suppressed TNFalpha-induced VCAM-1 mRNA expression. These actions of 17-epiestriol and genistein were significantly attenuated in the presence of the estrogen receptor antagonist ICI-182780. Other estrogenic compounds such as ethinyl estradiol and estrone did not have any effect on TNFalpha-induced VCAM-1 expression at the concentrations tested. We further show that, 1) 17-epiestriol induces the expression of endothelial nitric-oxide synthase mRNA and protein, 2) 17-epiestriol prevents TNFalpha-induced migration of NFkappaB into the nucleus, 3) N(G)-nitro-l-arginine methyl ester, an inhibitor of NO synthesis, abolishes 17-epiestriol-mediated inhibition of TNFalpha-induced VCAM-1 expression and migration of NFkappaB from the cytoplasm to the nucleus. Our results indicate that 17-epiestriol is more potent than 17-beta E(2) in suppressing TNFalpha-induced VCAM-1 expression and that this action is modulated at least in part through NO.     

Shoot organogenesis and mass propagation of Coleus forskohlii from leaf derived callus

A high frequency shoot organogenesis and plant establishment protocol has been developed for Coleus forskohlii from leaf derived callus. Optimal callus was developed from mature leaves on Murashige and Skoog (MS) medium supplemented with 2.4 μM kinetin alone. Shoots were regenerated from the callus on MS medium supplemented with 4.6 μM kinetin and 0.54 μM 1-naphthalene acetic acid. The highest rate of shoot multiplication was achieved at the sixth subculture and more than 150 shoots were produced per callus clump. Regenerated shootlets were rooted spontaneously on half-strength MS medium devoid of growth regulators. The in vitro raised plants were established successfully in soil. The amount of forskolin in in vitroraised plants and wild plants was estimated and found that they produce comparable quantity of forskolin. This in vitro propagation protocol should be useful for conservation as well as mass propagation of this plant. This revised version was published online in June 2006 with corrections to the Cover Date.     

Comparative effectiveness of an integrated pest management system and other control tactics for managing the spider mite Tetranychus ludeni (Acari: Tetranychidae) on eggplant

The effect of an integrated pest management (IPM) package, host plantresistance, Chrysoperla carnea predation and neem oil wereevaluated against the spider mite Tetranychus ludeni oneggplant(Solanum melongena L.) fields in 1996 and 1997, byestimating the mite population density and yield levels. Compared with the IPMpackage (Panruti local, C. carnea plus neem oil), thestandard (susceptible) eggplant variety (MDU₁) grown by farmers andtreated with an acaricide (dicofol) had significantly higher mite densities.Thepredator C. carnea was recorded in significantly lowernumbers in plots with the standard variety compared to a resistant variety(panruti local) with the full IPM package. Eggplant yield level and crop valuewere highest in the IPM-treated plots followed by Panruti local plusC. carnea. The standard variety treated with an acaricidehad the lowest yield and value. These results indicated the usefulness of hostplant resistance complemented by biorational control agents, such asC. carnea and neem oil, that these are suitable componentsin an IPM programme for managing the spider mite in endemic areas.     

Alpha-B crystallin gene (CRYAB) mutation causes dominant congenital posterior polar cataract in humans

Congenital cataracts are an important cause of bilateral visual impairment in infants. In a four-generation family of English descent, we mapped dominant congenital posterior polar cataract to chromosome 11q22-q22.3. The maximum LOD score, 3.92 at recombination fraction 0, was obtained for marker D11S898, near the gene that encodes crystallin alpha-B protein (CRYAB). By sequencing the coding regions of CRYAB, we found in exon 3 a deletion mutation, 450delA, that is associated with cataract in this family. The mutation resulted in a frameshift in codon 150 and produced an aberrant protein consisting of 184 residues. This is the first report of a mutation, in this gene, resulting in isolated congenital cataract.