Genetic Screening Identifies Cyanogenesis-Deficient Mutants of Lotus japonicus and Reveals Enzymatic Specificity in Hydroxynitrile Glucoside Metabolism

Cyanogenesis, the release of hydrogen cyanide from damaged plant tissues, involves the enzymatic degradation of amino acid–derived cyanogenic glucosides (α-hydroxynitrile glucosides) by specific β-glucosidases. Release of cyanide functions as a defense mechanism against generalist herbivores. We developed a high-throughput screening method and used it to identify cyanogenesis deficient (cyd) mutants in the model legume Lotus japonicus. Mutants in both biosynthesis and catabolism of cyanogenic glucosides were isolated and classified following metabolic profiling of cyanogenic glucoside content. L. japonicus produces two cyanogenic glucosides: linamarin (derived from Val) and lotaustralin (derived from Ile). Their biosynthesis may involve the same set of enzymes for both amino acid precursors. However, in one class of mutants, accumulation of lotaustralin and linamarin was uncoupled. Catabolic mutants could be placed in two complementation groups, one of which, cyd2, encoded the β-glucosidase BGD2. Despite the identification of nine independent cyd2 alleles, no mutants involving the gene encoding a closely related β-glucosidase, BGD4, were identified. This indicated that BGD4 plays no role in cyanogenesis in L. japonicus in vivo. Biochemical analysis confirmed that BGD4 cannot hydrolyze linamarin or lotaustralin and in L. japonicus is specific for breakdown of related hydroxynitrile glucosides, such as rhodiocyanoside A. By contrast, BGD2 can hydrolyze both cyanogenic glucosides and rhodiocyanosides. Our genetic analysis demonstrated specificity in the catabolic pathways for hydroxynitrile glucosides and implied specificity in their biosynthetic pathways as well. In addition, it has provided important tools for elucidating and potentially modifying cyanogenesis pathways in plants.     

Radiation synthesis, characterization and dye adsorption of alginate-organophilic montmorillonite nanocomposite

In this paper, the preparation, characterization and dye adsorption properties of nanocomposite (calcium alginate/organophilic montmorillonite) (CA/OMMT) were investigated. A new nanocomposite consisting of alginate and OMMT was prepared by polymerization using γ-rays irradiation as initiator. Physical characteristics of CA/OMMT were studied using X-ray diffraction (XRD), infrared spectrophotometery (IR), thermal gravimetric analysis (TGA), transmission electron microscopy (TEM) and the corresponding selected area electron diffraction (SAED). Two textile dyes, acid green B and direct pink 3B, were used as model anionic dye. Factors affecting dye sorption, such as pH, sorbent concentration and temperature of each dye solution were extensively investigated. It was found from the study that the sorption of dyes by the nanocomposite is pH-dependent and maximum sorption was obtained at pH 2. The thermodynamic data showed that dye adsorption onto alginate was spontaneous, exothermic, and a physisorption reaction. On the basis of the data of the present investigation, one could conclude that the as-prepared adsorbents exhibited excellent affinity for the dye, and can be applied to treat wastewater containing anionic dyes.     

Pharmacokinetic, metabolic, and antidiarrheal properties of ( and ) heptapeptides of sorbin in rodent

The C-terminal heptapeptide-amide (C7-sorbin) is the minimal biologically active fragment of sorbin inducing an increase in intestinal hydroelectrolytic absorption. An analogue (D7-sorbin), characterized by the replacement of the ultimate C-terminal amino acid -alanine-amide by -alanine-amide, was synthetized. For pharmacokinetic studies, D7-sorbin and C7-sorbin were tritium labeled. After IV injection, clearances were 10.6 and 30.2 ml⁻¹ for D7-sorbin and C7-sorbin, respectively, and MRT were 34 and 18 min. After SC administration, C_max attained 0.41% and 0.12% of the dose/ml, respectively. The IP route showed a 45-min delay before C_max and a 100% bioavailability for both peptides. D7-sorbin was principally excreted in urine, as shown by balance study, and in part in intact form, as controlled by mass spectrometry. D7-sorbin induced a significant decrease of the VIP-induced ileal secretion, previously observed with C7-sorbin. The change of -Ala to -Ala increased the stability of the synthetic C-terminal peptide of sorbin whereas its biological activity, bioavailability, and route of elimination were unchanged.     

Radiopacity additives in silicone stent materials reduce in vitro bacterial adherence

In vitro adherence of the nosocomial pathogensPseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, Serratia marcescens, andCandida albicans to select radiopaque silicone compounds was lower than that observed for the base silicone (p<0.03). Except forE. coli ATCC 11775 andCandida albicans, all microorganisms showed significantly lower adherence to a silicone compound impregnated with tantalum in comparison with a silicone compound impregnated with barium sulfate (p<0.05). Surface hydrophobicity of the silicone compounds did not show a direct correlation with the concentration of radiopacity additives or with degree of bacterial adherence. Scatchard analyses of data indicated that the number of adherence sites forP. aeruginosa on the base silicone, BaSO₄-silicone, and Ta-silicone were 9.2×10⁶ per mm², 6.1×10⁶ per mm², and 3.7×10⁶ per mm² respectively. As determined by the Langmuir adsorption isotherm, the dissociation constants for adheredP. aeruginosa to the base silicone, BaSO₄-silicone, and Ta-silicone were 2.50×10³ mm⁴, 1.45×10³ mm⁴, and 6.27×10³ mm⁴ respectively.Pseudomonas aeruginosa demonstrated first order kinetics of adherence to the silicone compounds with a half saturation time of 4.15 h for the base silicone, 1.06 h for the BaSO₄-silicone, and 2.14 h for the Ta-silicone. The use of Ta-silicone stents may delay the development of ascending urinary tract infections.