A peptide-mediated and hydroxyl radical HO*-involved oxidative degradation of cellulose by brown-rot fungi

A special low-molecular-weight peptide named Gt factor, was isolated and purified from the extracellular culture of brown-rot fungi Gloeophyllum trabeum via gel filtration chromatography and HPLC. It has been shown to reduce Fe³⁺ to Fe²⁺. Electron paramagnetic resonance (EPR) spectroscopy revealed Gt factor was able to drive H₂O₂ generation via a superoxide anion O₂ ^.- intermediate and mediate the formation of hydroxyl radical HO^. in the presence of O₂. All the results indicated that Gt factor could oxidize the cellulose, disrupt the inter- and intrahydrogen bonds in cellulose chains by a HO^. -involved mechanism. This resulted in depolymerization of the cellulose, which made it accessible for further enzymatic hydrolysis.     

HIV-2: the forgotten AIDS virus

HIV type 2 (HIV-2), a closely related retrovirus discovered a few years after HIV type 1, causes AIDS in only a minority of infected individuals. Determining why HIV-2 causes asymptomatic infection in most patients could further our understanding of HIV immunopathogenesis. Studies to date have suggested that both enhanced immune responses and lower viral replication could play a role. We summarize the important findings to date and highlight areas that warrant further exploration.     

Modulation of the antioxidant activity of HO* scavengers by albumin binding: a 19F-NMR study

The interaction between different HO(z.rad;) radical scavengers in a three-component antioxidant system has been investigated by means of 19F-NMR spectroscopy. This system is composed of bovine serum albumin (BSA), trolox, and N-(4-hydroxyphenyl)-trifluoroacetamide (CF(3)PAF). The antioxidant capacity of BSA and trolox has been assessed by measuring the amount of trifluoroacetamide (TFAM) arising from the radical mediated decomposition of CF(3)PAF. When assayed separately, both trolox and BSA behaved as antioxidants, as they were effective to protect CF(3)PAF from HO* radical-mediated decomposition. By contrast, trolox enhanced the production of TFAM in the presence of BSA, thus behaving as a pro-oxidant. Urate, carnosine, glucose, and propylgallate showed antioxidant properties both with or without BSA. CF(3)PAF and trolox were found to bind to BSA with association constants in the order of 5 x 10(3)M(-1) and to compete for the same binding sites. These results have been discussed in terms of BSA-catalysed cross-reactions between trolox-derived secondary radicals and CF(3)PAF.     

Damage to human erythrocytes by radiation-generated HO* radicals: molecular changes in erythrocyte membranes

The effectiveness of radiation-generated HO

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radicals in initiating erythrocyte hemolysis in the presence of oxygen and under anaerobic conditions and prehemolytic structural changes in the plasma-erythrocyte membrane were studied. Under anaerobic conditions the efficacy of HO

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radicals in induction of hemolysis was 16-fold lower than under air. In both conditions, hemolysis was the final consequence of changes of the erythrocyte membrane. Preceding hemolysis, the dominating process under anaerobic conditions was the aggregation of membrane proteins. The aggregates were principally formed by -S-S- bridges. A decrease in spectrin and protein of band 3 content suggests their participation in the formation of the aggregates. These processes were accompanied by changes in protein conformation determined by means of 4-maleimido-2,2,6,6-tetramethylpiperidine-N-oxyl (MSL) spin label attached to membrane proteins. Under anaerobic conditions, in the range of prehemolytical doses, the reaction of HO

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with lipids caused a slight (10-16%) increase in fluidity of the lipid bilayer in its hydrophobic region with a lack of lipid peroxidation. However, in the presence of oxygen, hemolysis was preceded by intense lipid peroxidation and by profound changes in the conformation of membrane proteins. At the radiation dose that normally initiates hemolysis a slight aggregation of proteins was observed. Changes were not observed in particular protein fractions. It can be suggested the cross-linking induced by HO

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radicals under anaerobic conditions and a lack of lipid peroxidation are the cause of a decrease in erythrocyte sensitivity to hemolysis. Contrary, under aerobic conditions, molecular oxygen suppresses cross-linking, catalysing further steps of protein and lipid oxidation, which accelerate hemolysis.     

Evaluation of N-hydroxy-2-thiopyridone as a nonmetal dependent source of the hydroxyl radical (HO.) in aqueous systems.

N-hydroxy-2-thiopyridone (1), an established source of the hydroxyl radical (HO., Boivin, J., Crepon, E., and Zard, S. Z. (1990) Tetrahedron Lett. 31, 6869-6872), produced HO. under conditions directly applicable to biological studies. Generation of HO. by subjecting 1 to irradiation with visible light was monitored in the following "HO." assays: deoxyribose degradation, addition to dimethyl sulfoxide, and hydroxylation of salicylate and phenol. All four assays demonstrated the production of HO. from 1 (added as a sodium salt) under mild conditions in aqueous buffer systems. An improved analysis method was developed for the phenol assay. A time course analysis demonstrated that a flux of HO. is generated from 1 throughout the irradiation period, in contrast to the classical Fenton reaction of H2O2 with a transition metal in which a burst of HO. is generated in a short time period. While a thiyl radical is generated from 1 concurrent with HO. generation, this species does not contribute to, or interfere with, any of the HO. assays, suggesting that it is weakly reactive in aqueous buffers. Thus, irradiation of 1 can be used as an alternative, complementary, approach for the unequivocal generation of the biologically significant and reactive HO..     

Reevaluation of the reactivity of hydroxylamine with O2-/HO2

The reactivity of hydroxylamine with HO2/O2- radicals was studied by pulse radiolysis and stopped-flow photolysis over a pH range of 1.1-10.5. Upper limits for the rate of reaction indicate that hydroxylamine, if it reacts at all, reacts at a very slow rate. Its use as an indicator for O-2 and an assay for superoxide dismutase is, therefore, inappropriate.     

Micro-invertebrates conservation: forgotten biodiversity

The concern about the preservation of biodiversity is due, in part, to a great level of media coverage granted in the last few years to global warming and consequential climatic changes. However, there are still considerably large gaps in scientific knowledge regarding the ecological status of many species, which results in an absence of conservation strategy for most of Earth’s biodiversity in need of it. The extinction of many animal and plant species can have catastrophic consequences on the ecosystems’ balance and also in human well-being, resultant from the break of ecological services. To exemplify how a specific group of microscopic animals can be endangered, I have analyzed the case of the phylum Tardigrada. Tardigrades are microscopic animals that inhabit most environments: terrestrial, freshwater and marine. Even though many species are widespread and the terrestrial ones granted with cryptobiotic skills, they are adapted to each habitat type and, additionally, to local environmental patterns. This means that these tiny metazoans can be under significant environmental pressure in the various habitat types they are found in. The potential need of protective and compensatory measures aiming for appropriate conservation of these life forms is discussed, as is the need of studying for their objective elaboration.     

Evidence against the 1:2:2:1 quartet DMPO spectrum as the radical adduct of the lipid alkoxyl radical.

It was reported that the electron paramagnetic resonance (EPR) spectrum of 5,5-dimethyl-1-pyrroline N-oxide (DMPO)/lipid alkoxyl radical exhibited a quartet with 1:2:2:1 relative intensity that is identical to that of DMPO/hydroxyl radical (K. M. Schaich and D. C. Borg, 1990, Free Radicals Res. Commun. 9, 267-278). We repeated these EPR experiments using HPLC separation of radical adducts and isotope substitution. We found that the HPLC/EPR chromatogram of the radical adduct with a 1:2:2:1 quartet obtained by the reduction of methyl linoleate hydroperoxide (MLOOH) with Fe2+ exhibited identical retention time to that of the DMPO/OH radical adduct obtained from the Fenton reaction in two different solvent systems. Upon performing the same reaction in 17O-enriched water, the 17O-hyperfine coupling constants due to DMPO/17OH were identified. Ultimately, approximately 80-90% of the total DMPO/OH is derived from water by an iron-dependent nucleophilic addition reaction. Initially, a water-independent mechanism also significantly contributes to DMPO/OH formation. Regardless of its mechanism of formation, the 1:2:2:1 quartet radical adduct of DMPO formed during the reduction of MLOOH by Fe2+ is in fact DMPO/OH.     

Necrobacillosis: a forgotten disease.

Over four years five previously healthy young adults developed necrobacillosis , a severe septicaemic illness caused by Fusobacterium necrophorum. The infections were characterised by sore throat followed by rigors and the formation of metastatic abscesses and all caused considerable diagnostic confusion.     

Microtubules: forgotten players in the apoptotic execution phase

A cell entering the execution phase of apoptosis (regulated cell death) undergoes characteristic rearrangements, in which the cytoskeleton has major roles. Historically, this reorganisation has been attributed entirely to actomyosin contractility, with microtubule and intermediate filament systems both reported to be lost at an early stage. However, recent results indicate that microtubule networks re-form during the later stages of apoptosis and assist in the dispersal of nuclear and cellular fragments--steps that are thought to be important for preventing inflammation. Here, we discuss the roles of the cytoskeleton during apoptosis and challenge current thinking that actin is the sole functional component driving all major execution phase events.