<Emphasis Type="Italic">In vitro</Emphasis> organogenesis and antioxidant enzymes activity in <Emphasis Type="Italic">Acanthophyllum sordidum</Emphasis>

The effect of various hormonal combinations on callus formation and regeneration of shoot and root from leaf derived callus of Acanthophyllum sordidum Bunge ex Boiss. has been studied. Proteins and activity of antioxidant enzymes were also evaluated during shoot and root organogenesis from callus. Calli were induced from leaf explants excised from 30-d-old seedlings grown on Murashige and Skoog medium containing 4.52 μM 2,4-dichlorophenoxyacetic acid + 4.65 μM kinetin. Maximum growth of calli and the most efficient regeneration of shoots and roots occurred with 2.69 μM 1-naphthalene acetic acid (NAA), 2.69 μM NAA + 4.54 μM thidiazuron and 2.46 μM indole-3-butyric acid. Protein content decreased in calli and increased significantly during regeneration of shoots from callus. Superoxide dismutase activity decreased in calli comparing to that of seedlings, then increased in regenerated shoots and roots. High catalase activity was detected in seedlings and regenerated shoots, whereas high peroxidase activity was observed in calli and regenerated roots.     

辣根过氧化物酶的热稳定剂

保持酶的天然状态和高催化特性具有重要的意义。本研究筛选了辣根过氧化物酶(HRP)的稳定剂并研究了其作用机制。结果发现硫酸镁和明胶能够显著提高HRP的热稳定性,并且两者具有协同作用。在硫酸镁和明胶组成的酶稳定剂存在的条件下,HRP在50oC保温80h后仍能保持89%的活性,常温下存放90d后可保持57%的活性,而未加稳定剂的对照样品中HRP的残留活性分别为6%和小于1%。通过对HRP的Soret带吸收光谱,色氨酸内源荧光,ANS荧光进行分析,揭示酶稳定剂可以明显降低在加热条件下HRP的变性程度,从而维持较为稳定的天然构象。     

Vital roles of an interhelical insertion in catalase-peroxidase bifunctionality

The loop connecting the F and G helices of catalase-peroxidases contains a approximately 35 amino acid structure (the FG insertion) that is absent from monofunctional peroxidases. These two groups of enzymes share highly similar active sites, yet the monofunctional peroxidases lack appreciable catalase activity. Thus, the FG insertion may serve a role in catalase-peroxidase bifunctionality, despite its peripheral location relative to the active site. We produced a variant of Escherichia coli catalase-peroxidase (KatG) lacking its FG insertion (KatG(DeltaFG)). Absorption spectra indicated the heme environment of KatG(DeltaFG) was highly similar to wild-type KatG, but the variant retained only 0.2% catalase activity. In contrast, the deletion reduced peroxidase activity by only 50%. Kinetic parameters for the peroxidase and residual catalase activities of KatG(DeltaFG) as well as pH dependence studies suggested that the FG insertion supports hydrogen-bonded networks critical for reactions involving H2O2. The structure also appears to regulate access of electron donors to the active site.     

Salicylic acid and methyl jasmonate improve chilling tolerance in cold-stored lemon fruit (Citrus limon)

Chilling injury (CI) is associated with the degradation of membrane integrity which can be aligned to phenolic oxidation activated by polyphenol oxidase (PPO) and peroxidase (POD), enzymes responsible for tissue browning. Phenylalanine ammonia-lyase (PAL) is a further enzyme prominent in the phenolic metabolism that is involved in acclimation against chilling stress. It was hypothesized that treatment with methyl jasmonate (MJ) and salicylic acid (SA) may enhance chilling tolerance in lemon fruit by increasing the synthesis of total phenolics and PAL by activating the key enzyme regulating the shikimic acid pathway whilst inhibiting the activity of POD and PPO. Lemon fruit were treated with 10 μM MJ, 2 mM SA or 10 μM MJ plus 2 mM SA, waxed, stored at −0.5, 2 or 4.5 °C for up to 28 days plus 7 days at 23 °C. Membrane integrity was studied by investigating membrane permeability and the degree of membrane lipid peroxidation in lemon flavedo following cold storage. The 10 μM MJ plus 2 mM SA treatment was most effective in enhancing chilling tolerance of lemon fruit, significantly reducing chilling-induced membrane permeability and membrane lipid peroxidation of lemon flavedo tissue. This treatment also increased total phenolics and PAL activity in such tissue while inhibiting POD activity, the latter possibly contributing to the delay of CI manifestation. PPO activity was found to be a poor biochemical marker of CI. Treatment with 10 μM MJ plus 2 mM SA resulted in an alteration of the phenolic metabolism, enhancing chilling tolerance, possibly through increased production of total phenolics and the activation of PAL and inhibition of POD.     

An 18 kDa Acid Phosphatase from Chicken Heart Possesses Phosphotransferase Activity

A low molecular weight acid phosphatase was purified to homogeneity from chicken heart with a specific activity of 42 U/mg and a recovery of about 1%. Nearly 800 fold purification was achieved. The molecular weight was estimated to be 18 kDa by SDS-polyacrylamide gel electrophoresis. Para-nitrophenyl phosphate, phenyl phosphate and flavin mononucleotide were efficiently hydrolysed by the enzyme and found to be good substrates. Fluoride and tartrate had no inhibitory effect while phosphate, vanadate and molybdate strongly inhibited the enzyme. The acid phosphatase was stimulated in the presence of glycerol, ethylene glycol, methanol, ethanol and acetone, which reflected the phosphotransferase activity. When phosphate acceptors such as ethylene glycol concentrations were increased, the ratio of phosphate transfer to hydrolysis was also increased, demonstrating the presence of a transphosphorylation reaction where an acceptor can compete with water in the rate limiting step involving hydrolysis of a covalent phospho enzyme intermediate. Partition experiments carried out with two substrates, para-nitrophenyl phosphate and phenyl phosphate, revealed a constant product ratio of 1.7 for phosphotransfer to ethylene glycol versus hydrolysis, strongly supporting the existence of common covalent phospho enzyme intermediate. A constant ratio of K _cat/K _m, 4.3×10⁴, found at different ethylene glycol concentrations, also supported the idea that the rate limiting step was the hydrolysis of the phospho enzyme intermediate.     

A chromium-tolerant plant growing in Cr-contaminated land

In this paper, a kind of Typhaceae plant species, Typha angustifolia L, with a high tolerance to Cr is described. Experiments were carried out to examine its ability to tolerant Cr and its physiological response. The results showed that there was no difference in growth, plant height, and biomass response to external Cr (VI) between the plants exposed to 100 microM Cr (VI) and control (0 microM), while increasing Cr levels to 200-800 microM induced a significant decrease in plant height and biomass, but no significant injury was detected, even for the plants exposed to 800 microM Cr. Chromium induced significant increases in superoxide dismutase (SOD) and peroxidase (POD) activities. Meanwhile, a significantly positive correlation was found between Cr and Mn or Cu in leaves and roots, respectively. The Cr tolerance of the plant appeared to be associated with the enhancement of SOD and POD activities and the improvement in uptake and translocation of the essential microelements.     

Drought and Oxidative Load in the Leaves of C3 Plants: a Predominant Role for Photorespiration?

Although active oxygen species are produced at high rates inboth the chloroplasts and peroxisomes of the leaves of C₃ plants,most attention has focused on the potentially damaging consequencesof enhanced chloroplastic production in stress conditions suchas drought. This article attempts to provide quantitative estimatesof the relative contributions of the chloroplast electron transportchain and the glycolate oxidase reaction to the oxidative loadplaced on the photosynthetic leaf cell. Rates of photorespiratoryH₂O₂ production were obtained from photosynthetic and photorespiratoryflux rates, derived from steady-state leaf gas exchange measurementsat varying irradiance and ambient CO₂. Assuming a 10 % allocationof photosynthetic electron flow to the Mehler reaction, photorespiratoryH₂O₂ production would account for about 70 % of total H₂O₂ formedat all irradiances measured. When chloroplastic CO₂ concentrationrates are decreased, photorespiration becomes even more predominantin H₂O₂ generation. At the increased flux through photorespirationobserved at lower ambient CO₂, the Mehler reaction would haveto account for more than 35 % of the total photosynthetic electronflow in order to match the rate of peroxisomal H₂O₂ production.The potential signalling role of H₂O₂ produced in the peroxisomesis emphasized, and it is demonstrated that photorespiratoryH₂O₂ can perturb the redox states of leaf antioxidant pools.We discuss the interactions between oxidants, antioxidants andredox changes leading to modified gene expression, particularlyin relation to drought, and call attention to the potentialsignificance of photorespiratory H₂O₂ in signalling and acclimation.     

Selenium status of patients with Balkan endemic nephropathy

The selenium (Se) contents in common cereals in endemic and nonendemic areas in Serbia are very low. Plasma Se levels of both patients and healthy subjects, were also low, reflecting low Se intakes. Patients with Balkan endemic nephropathy (BEN) had significantly lower (p<0.05) plasma Se levels than healthy individuals, both from regions close to endemic areas, and from Belgrade. Mean plasma Se of BEN patients was slightly but insignificantly higher in samples taken immediately after dialysis than in those taken before, suggesting that very little of the Se present in plasma is dialyzable. Plasma SeGSH-Px activities before and after hemodialysis in both BEN and Nonendemic chronic renal failure (NCRF) patients were not significantly different, but BEN patients had lower enzyme activities than those with NCRF and healthy controls. In BEN patients, a significant correlation between plasma Se and SeGSH-Px activity was found. NCFR patients were with diagnoses: TBC of kidneys, chronic glomerulonephritis, chronic pyelonephritis, and polycystic kidneys.     

Neisseria gonorrhoeae possesses two nicotinamide adenine dinucleotide-independent lactate dehydrogenases

Abstract An important metabolic capability of Neisseria gonorrhoeae is the utilization of host-derived lactate. Two isoenzymes of the membrane-associated, pyridine dinucleotide-independent type of lactate dehydrogenase (iLDH) participate in lactate assimilation, but exhibit distinctive properties. Isoenzyme iLDH-I utilized lactate exclusively as substrate, exhibiting a preference for the D-isomer. In contrast, isoenzyme iLDH-II exhibited broad substrate specificity (lactate, phenyllactate, and 4-hydroxyphenyllactate), but was stereospecific for the L-isomers. These results explain the difficulty in isolating mutants unable to utilize lactate.