Copper- and arsenate-induced oxidative stress in Holcus lanatus L. clones with differential sensitivity

The biochemical responses of Holcus lanatus L. to copper and arsenate exposure were investigated in arsenate‐tolerant and ‐non‐tolerant plants from uncontaminated and arsenic/copper‐contaminated sites. Increases in lipid peroxidation, superoxide dismutase (SOD) activity and phytochelatin (PC) production were correlated with increasing copper and arsenate exposure. In addition, significant differences in biochemical responses were observed between arsenate‐tolerant and ‐non‐tolerant plants. Copper and arsenate exposure led to the production of reactive oxygen species, resulting in significant lipid peroxidation in non‐tolerant plants. However, SOD activity was suppressed upon metal exposure, possibly due to interference with metallo‐enzymes. It was concluded that in non‐tolerant plants, rapid arsenate influx resulted in PC production, glutathione depletion and lipid peroxidation. This process would also occur in tolerant plants, but by decreasing the rate of influx, they were able to maintain their constitutive functions, detoxify the metals though PC production and quench reactive oxygen species by SOD activity.     

Probing the Structural Basis for Enzyme-Substrate Recognition In Cu,Zn Superoxide Dismutase

A full understanding of enzyme-substrate interactions requires a detailed knowledge of their structural basis at atomic resolution. Crystallographic and biochemical data have been analyzed with coupled computational and computer graphic approaches to characterize the molecular basis for recognition of the superoxide anion substrate by Cu. Zn superoxide dismutase (SOD). Detailed analysis of the bovine SOD structure aligned with SOD sequences from 15 species provides new results concerning the significance and molecular basis for sequence conservation. Specific roles have been assigned for all 23 invariant residues and additional residues exhibiting functional equivalence. Sequence invariance is dominated by 15 residues that form the active site stcreochemistry. supporting a primary biological function of superoxide dismutation. Using data from crystallographic structures and site-directed mutants, we are testing the role of individual residues in the active site channel, including (in human SOD) Glu132, Glu133, Lys136, Thr137, and Arg 143. Electrostatic calculations incorporating molecular flexibility suggest that the region of positive electrostatic potential in and over the active site channel above the Cu ion sweeps through space during molecular motion to enhance the facilitated diffusion responsible for the enzyme's rapid catalytic rate.     

The effects of deuterium oxide (2H2O) on the polymerization of tubulin in vitro

The initial rate and final extent of polymerization of both bovine brain tubulin and sea urchin egg tubulin were enhanced in the presence of ²H₂O. The yields were increased in association with the elevation of the ²H₂O concentration. ²H₂O also reduced the critical concentration for polymerization of brain tubulin. Thermodynamic analysis was attempted using the temperature dependence of the critical concentration for polymerization in the presence of ²H₂O. We obtained linear van 't Hoff plots and calculated thermodynamic parameters which were positive and were increased with the elevation of the ²H₂O concentration. The enhancement of the polymerization of tubulin by ²H₂O could, therefore, be the result of the strenghening of intra-and/or inter-molecular hydrophobic interactions of the tubulin molecules. We believe that the increase in lenghth and number of microtubules of the mitotic spindles in the dividing cells of the eukaryotes with ²H₂O may be caused by the direct involvement of ²H₂O in the polymerization of tubulin.     

Rare earth thiocyanate complexes with tripiperidinophosphine oxide: synthesis and characterization. Crystal structure of Pr(SCN)3(tpppO)3

The synthesis and characterization of rare-earth (La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu and Y) thiocyanate adducts with tripiperidinophosphine oxide (tpppO) with general formula (RE)(SCN)₃(tpppO)₃ are reported. Conductance measurements in acetonitrile indicate the non-electrolytic nature of the complexes. Infrared absorption spectra evidence that the SCN⁻ ion coordinates through the nitrogen atom (isothiocyanate form) and that tpppO coordinates through the phosphoryl oxygen. X-ray powder patterns suggest the existence of three different crystal forms: (1) La; (2) an isomorphous series including Ce, Nd and Pr; and (3) another isomorphous series, including Sm, Gd, Eu, Ho, Er, Tb, Lu and Y. The visible spectra of the Nd adduct and the calculated parameters β = 0.98, b^1/2 = 0.072 and δ = 1.06 indicate that the metal-ligand bonds are essentially electrostactic. The emission spectra of the Eu compound showed ⁵D₀ → ⁷F_J bands (J = 0, 1, 2), suggesting a C_3v symmetry for the coordination polyhedron. The lifetime of the ⁵D₀ state is 1.28 ms. The emission spectra of the Tb complex presented ⁵D₄ → ⁷F_J bands (J = 4, 5, 6) and the Dy complex showed the ⁴F_9/2 → ⁶H_13/2 band. The structure of the Pr complex showed that the coordination polyhedron is a trigonal antiprism, with the isothiocyanate anions in one base and three tpppO ligands in the other. Thermal analyses (TG-DTG) were carried out for the Ce, Nd and Gd adducts. Mass losses start between 250 and 334 °C. The final residues at 1300 °C are the corresponding phosphates.     

Periplasmic superoxide dismutases in Aquaspirillum magnetotacticum

Aquaspirillum magnetotacticum MS-1 cells cultured microaerobically (dissolved O₂ tension 1% of saturation), expressed proteins with superoxide dismutase (SOD) activity. The majority (roughly 95%) of total cell superoxide dismutase activity was located in the cell periplasm with little or no activity in the cell cytoplasm. Irontype SOD (FeSOD) contributed 88% of the total activity activity detected, although a manganese-type SOD (MnSOD) was present in the periplasm as well. Cells cultured at a higher dissolved O₂ tension (10% of saturation) expressed increased activity of the MnSOD relative to that of the FeSOD.     

Short-chain fatty acids and polyamines in the pathogenesis of necrotizing enterocolitis: Kinetics aspects in gnotobiotic quails

Despite years of investigation, pathogenesis of necrotizing enterocolitis (NEC) remains elusive. Bacterial metabolites were implicated by several authors but their roles remain controversial. The aim of our study was to investigate the role of SCFAs and polyamines through a kinetic study of histological and macroscopical digestive lesions in monobiotic quails. Germ-free quails, inoculated with a Clostridium butyricum strain involved in a NEC case, were fed or not with a diet including lactose (7%). Quails were sacrificed at various times between D7 and D24 after bacterial inoculation. NEC-like lesions, i.e. thickening, pneumatosis, and hemorrhages, occurred only in lactose-fed quails and increased with time. The main histological characteristics were infiltrates of mononuclear cells, then heterophilic cells, then gas cyst and necrosis. The first event observed, before histological and macroscopical lesions, is a high production of butyric acid, which precedes an increase of iNOS gene expression. No difference in polyamines contents depending on the diet was observed. These results show the major role of butyric acid produced by commensal bacteria in the onset of the digestive lesions.     

Sustained delivery of exogenous melatonin influences biomarkers of oxidative stress and total antioxidant capacity in summer-stressed anestrous water buffalo (Bubalus bubalis)

High ambient temperature during summer in tropical and subtropical countries predisposes water buffaloes (Bubalus bubalis) to develop oxidative stress having antigonadotropic and antisteroidogenic actions. Melatonin is a regulator of seasonal reproduction in photoperiodic species and highly effective antioxidant and free radical scavenger. Therefore, a study was designed to evaluate the effect of sustained-release melatonin on biomarkers of oxidative stress i.e., the serum malondialdehyde (MDA) and nitric oxide (NO), and the total antioxidant capacity (TAC). For the study, postpartum buffaloes diagnosed as summer anestrus (absence of overt signs of estrus, concurrent rectal examination, and RIA for serum progesterone) were grouped as treated (single subcutaneous injection of melatonin at 18 mg/50 kg body weight dissolved in sterilized corn oil as vehicle, n = 20) and untreated (subcutaneous sterilized corn oil, n = 8). Blood sampling for estimation of serum TAC and MDA (mmol/L) and NO (μmol/L) was carried out at 4 days of interval from 8 days before treatment till 28 days after treatment or for the ensuing entire cycle length. Results showed serum TAC concentration was higher in the treatment group with a significant (P < 0.05) increasing trend, whereas MDA and NO revealed a significant (P < 0.05) decline. Serum MDA and NO were higher in control compared with those of treatment group. Moreover, buffaloes in the treatment group showed 90% estrus induction with 18.06 ± 1.57 days mean interval from treatment to the onset of estrus. These results report that melatonin has a protective effect by elevating antioxidant status and reducing oxidative stress resulting in the induction of cyclicity in summer-stressed anestrous buffaloes.     

Screening of thermotolerant yeasts as producers of superoxide dismutase

Abstract A screening procedure for highly thermostable yeast superoxide dismutase was developed. Growth yields at various temperatures were estimated for ten mesophilic and thermotolerant strains, belonging to the genera Saccharomyces, Kluyveromyces and Pichia . Higher yields at 45°C were obtained for K. lactis 90-3 and 90-4. A correlation between the ability to grow at higher temperature and the thermostability of the superoxide dismutase enzyme synthesized was observed. A comparison of the operational stability of the superoxide dismutase of all tested strains suggests that the enzyme of K. lactis strains was more thermostable than that of the other tested microorganisms.