Redox reactions associated with iron release from mammalian ferritin

The early redox events involved in iron reduction and mobilization in mammalian ferritin have been investigated by several techniques. Sedimentation velocity measurements of ferritin samples with altered core sizes, prepared by partial reduction and Fe2+ chelation, suggest two different events occur during iron loss from the ferritin core. Reductive optical titrations confirm this biphasic behavior by showing that the first 20-30% of core reduction has different optical properties than the latter 70-80%. Proton uptake during initial core reduction is near zero, but as the percent core reduction increases, the proton uptake (H+/e) values increase to 2 H+/e (2 H+/Fe3+ reduced) as core reduction approaches 1 e/Fe3+. Coulometric reduction of ferritin by mediators of different redox potential and different cross-sectional areas show a two-phase sigmoidal reaction pattern in which initial core reduction occurs at a slower rate than later core reduction. The above experiments were all conducted in the absence of iron chelators so that the observed results were all attributed to core reduction rather than the combined effects of core reduction accompanied by Fe2+ chelation. The coulometric reduction of ferritin by various mediators shows a correlation more with reduction potential than with molecular cross-sectional area. The role of the ferritin channels in core reduction is considered in terms of the reported results.     

Extracellular iron reduction is mediated in part by neutral red and hydrogenase in Escherichia coli

Both microbial iron reduction and microbial reduction of anodes in fuel cells can occur by way of soluble electron mediators. To test whether neutral red (NR) mediates iron reduction, as it does anode reduction, by Escherichia coli, ferrous iron levels were monitored in anaerobic cultures grown with amorphous iron oxide. Ferrous iron levels were 19.4 times higher in cultures fermenting pyruvate in the presence of NR than in the absence of NR. NR did not stimulate iron reduction in cultures respiring with nitrate. To explore the mechanism of NR-mediated iron reduction, cell extracts of E. coli were used. Cell extract-NADH-NR mixtures had an enzymatic iron reduction rate almost 15-fold higher than the chemical NR-mediated iron reduction rate observed in controls with no cell extract. Hydrogen was consumed during stationary phase (in which iron reduction was detectable) especially in cultures containing both NR and iron oxide. An E. coli hypE mutant, with no hydrogenase activity, was also impaired in NR-mediated iron reduction activity. NR-mediated iron reduction rates by cell extracts were 1.5 to 2 times higher with hydrogen or formate as the electron source than with NADH. Our findings suggest that hydrogenase donates electrons to NR for extracellular iron reduction. This process appears to be analogous to those of iron reduction by bacteria that use soluble electron mediators (e.g., humic acids and 2,6-anthraquinone disulfonate) and of anode reduction by bacteria using soluble mediators (e.g., NR and thionin) in microbial fuel cells.     

Transmembrane ferricyanide reduction by cells of the yeastSaccharomyces cerevisiae

Both respiratory-competent and respiratory-deficient yeast cells reduce external ferricyanide. The reduction is stimulated by ethanol and inhibited by the alcohol dehydrogenase inhibitor, pyrazole. The reduction of ferricyanide is not inhibited by inhibitors of mitochondrial or microsomal ferricyanide reduction. Cells in exponential-phase growth show a much higher rate of ferricyanide reduction. The reduction of ferricyanide is accompanied by increased release of protons by the yeast cells. We propose that the ferricyanide reduction is carried out by a transmembrane NADH dehydrogenase.     

Metabolic Interactions between Spinach Leaf Nitrite Reductase and Ferredoxin-NADP Reductase: Competition for Reduced Ferredoxin

Steady state rates of NADP reduction decline upon commencement of nitrite reduction in reconstituted chloroplast preparations. Similarly, steady state rates of nitrite reduction are lower, but not zero, during concurrent NADP reduction. These results imply that competition for substrate occurs and suggest that nitrite reduction can successfully compete for reduced ferredoxin, even at high rates of NADP reduction.     

Extracellular Iron Reduction Is Mediated in Part by Neutral Red and Hydrogenase in Escherichia coli

Both microbial iron reduction and microbial reduction of anodes in fuel cells can occur by way of soluble electron mediators. To test whether neutral red (NR) mediates iron reduction, as it does anode reduction, by Escherichia coli, ferrous iron levels were monitored in anaerobic cultures grown with amorphous iron oxide. Ferrous iron levels were 19.4 times higher in cultures fermenting pyruvate in the presence of NR than in the absence of NR. NR did not stimulate iron reduction in cultures respiring with nitrate. To explore the mechanism of NR-mediated iron reduction, cell extracts of E. coli were used. Cell extract-NADH-NR mixtures had an enzymatic iron reduction rate almost 15-fold higher than the chemical NR-mediated iron reduction rate observed in controls with no cell extract. Hydrogen was consumed during stationary phase (in which iron reduction was detectable) especially in cultures containing both NR and iron oxide. An E. coli hypE mutant, with no hydrogenase activity, was also impaired in NR-mediated iron reduction activity. NR-mediated iron reduction rates by cell extracts were 1.5 to 2 times higher with hydrogen or formate as the electron source than with NADH. Our findings suggest that hydrogenase donates electrons to NR for extracellular iron reduction. This process appears to be analogous to those of iron reduction by bacteria that use soluble electron mediators (e.g., humic acids and 2,6-anthraquinone disulfonate) and of anode reduction by bacteria using soluble mediators (e.g., NR and thionin) in microbial fuel cells.     

Triphasic reduction of cytochrome b and the protonmotive Q cycle pathway of electron transfer in the cytochrome bc1 complex of the mitochondrial respiratory chain

Reduction of cytochrome b in isolated succinate-cytochrome c reductase is a triphasic reaction. Initially, there is a relatively rapid, partial reduction of the cytochrome b, the rate of which matches the rate of reduction of cytochrome c1. This is followed by partial or complete reoxidation of the b, which is then followed by slow rereduction. At very low concentrations of succinate, the initial partial reduction of b is followed by reoxidation, but the third (rereduction) phase is absent, owing to insufficient substrate to rereduce the cytochromes. If antimycin is added at various times during the triphasic reaction, it inhibits the reoxidation and also inhibits the rereduction phase. Antimycin does not inhibit the initial phase of b reduction and, if added before or during this phase, it causes reduction of b to proceed to completion as a monophasic reaction. Myxothiazol inhibits the first phase of b reduction and the subsequent reoxidation, but does not inhibit the third, slow phase of b reduction. The resulting monophasic reduction of b which is observed in the presence of myxothiazol is slower than that in the presence of antimycin. The combination of both inhibitors, whether added together or successively during the triphasic reaction, completely inhibits b reduction. The triphasic reduction of cytochrome b is consistent with electron transfer by a protonmotive Q cycle in which there are two pathways for cytochrome b reduction. One pathway allows the initial phase of cytochrome b reduction by a myxothiazol-sensitive reaction in which reduction of b by ubisemiquinone is linked to reduction of iron-sulfur protein and cytochrome c1 by ubiquinol. In the second phase of the triphasic reaction, the b cytochromes are reoxidized by ubiquinone or ubisemiquinone through an antimycin-sensitive reaction. If oxidation of ubiquinol by iron-sulfur protein is blocked, either by myxothiazol or by reduction of iron-sulfur protein and cytochrome c1, the b cytochromes can be reduced by reversal of the antimycin-sensitive pathway, thus accounting for the third phase of b reduction.     

Reduction of perchlorate by an anaerobic enrichment culture

Summary A mixed bacterial culture capable of reducing perchlorate stoichiometrically to chloride under naerobic conditions was enriched from municipal digester sludge. The reduction of 10 mM perchlorate resulted in oxidation of the medium and cessation of perchlorate reduction. The activity was recovered on addition of a reducing agent. Addition of air to the culture during perchlorate reduction immediately terminated the process and aeration for 12 h permanently destroyed the ability of the culture to reduce perchlorate. The culture also reduced nitrite, nitrate, chlorite, chlorate and sulfate. The presence of 10 mM nitrite or chlorite completely inhibited perchlorate reduction, whereas the same concentration of chlorate decreased the reduction rate. Nitrate or sulfate did not affect perchlorate reduction. Chlorate and chlorite, suspected intermediates in the reduction of perchlorate to chloride, were not detected in any cultures during reduction of perchlorate.     

Energy metabolism in Saccharomyces cerevisiae. Effect of progressive starvation on respiration and pyridine nucleotide reduction linked to ethanol oxidation in intact cells

The progressive effects of aerobic starvation on endogenous and ethanol-linked respiration and pyridine nucleotide reduction have been studied in the yeast Saccharomyces cerevisiae. Three distinct phases of pyridine nucleotide reduction were observed when ethanol was added to unstarved yeast: an initial phase of rapid reduction and accelerating respiration (A); a steady-state phase of reduction with maximal respiration (B); a final phase of rapid reduction at anaerobiosis (C).During the first 5 hr of aeration, the steady-state Phase B was replaced by a phase of slow pyridine nucleotide reduction, while Phases A and C were unaffected. During this period, both endogenous pyridine nucleotide reduction and endogenous respiration decreased sharply.Between 5 and 22 hr of aeration, the endogenous level of reduced pyridine nucleotide declined further, while endogenous respiration remained unchanged. Concurrently, the extent of the Phase A reduction doubled.The addition of ethanol to aerobic, unstarved yeast stimulated a rapid pyridine nucleotide reduction, with further reduction occurring at anaerobiosis. Under anaerobic conditions, the addition of ethanol to unstarved yeast caused little further reduction of pyridine nucleotide. Two hours of starvation decreased the extent of the endogenously supported anaerobic reduction and correspondingly increased the ethanol-induced reduction. These results suggest that, in unstarved yeast, reducing equivalents derived from ethanol under aerobic conditions and those derived from endogenous carbohydrate under anaerobic conditions have access to the same pool of pyridine nucleotide. With starvation, this pool becomes accessible to ethanol-derived (or ethanol-mobilized) reducing equivalents under anaerobic conditions.     

Quinone interaction with the respiratory chain-linked NADH dehydrogenase of beef heart mitochondria. II. Duroquinone reductase activity

1. Enzymatic reduction of 2,3,5,6-tetramethyl-1,4-benzoquinone (duroquinone) by NADH can be used in an assay procedure for the NADH dehydrogenase. The reduction of this quinone occurs in the region of the electron transport system between the primary dehydrogenase and the cytochrome system as defined by the almost complete loss of reductase activity following piericidin A treatment.

2. Duroquinone reduction can be distinguished from ubiquinone 2 reduction by the marked inhibition of the former following phospholipase C, poly- -lysine, or chloroquine diphosphate treatment. In addition, duroquinone reduction requires the presence of endogenous ubiquinone 10 specifically whereas ubiquinone 2 reduction does not require the presence of endogenous quinone. These observations are consistent with the nonequivalency of the reduction sites of duroquinone and ubiquinone 2.

3. Duroquinol can be utilized as an electron donor for the energy-linked reduction, of NAD⁺. Duroquinol reduction of NAD⁺ is dependent upon the presence of ATP, is inhibited by oligomycin, carbonyl cyanide p-trifluoro methoxyphenylhydrazone and piericidin A, and is not inhibited by antimycin A at levels which inhibit electron transport.

4. Duroquinone reduction as well as ubiquinone 2 reduction are inhibited almost completely by phospholipase A, p-chloromercuribenzoate, o-phenanthroline, and Triton X100 treatments.     

Electrochemical reduction of methemoglobin either directly or with flavin mononucleotide as a mediator

Electrochemical reduction of methemoglobin on a platinum electrode is studied by means of thin layer spectroelectrochemistry. For methemoglobin alone in solution, direct reduction is very slow even for potentials close to those of the reduction of the solvent. The reduction of a methemoglobin-oxyhemoglobin mixture with an imposed potential causes the electrochemical reduction of oxygen, the conversion of oxyhemoglobin into deoxyhemoglobin, and a simultaneous transformation of part of the molecules into methemoglobin. When fixed oxygen has disappeared, reduction of methemoglobin takes place. The reduction of methemoglobin and deoxyhemoglobin is catalyzed by the presence of flavin mononucleotide (FMN). For the oxyhemoglobin-methemoglobin mixture, flavin makes a fast deoxygenation of oxyhemoglobin without a change in the oxidation state of the iron. It also allows the rapid reduction of methemoglobin. In each case, the resulting deoxyhemoglobin solutions do not show any electrolysis-induced modification of the equilibrium curves for oxygen binding.     

Chemical and biological mobilization of Fe(III) in marsh sediments

Iron reduction in marine sulfitic environments may occur via a mechanism involving direct bacterial reduction with the use of hydrogen as an electron donor, direct bacterial reduction involving carbon turnover, or by indirect reduction where sulfide acts to reduce iron. In the presented experiments, the relative importance of direct and indirect mechanisms of iron reduction, and the contribution of these two mechanisms to overall carbon turnover has been evaluated in two marsh environments. Sediments collected from two Northeastern US salt marshes each having different Fe (III) histories were incubated with the addition of reactive iron (as amorphous oxyhydroxide). These sediments were either incubated alone or in conjunction with sodium molybdate. Production of both inorganic and organic pore water constituents and a calculation of net carbon production were used as measures to compare the relative importance of direct bacterial reduction and indirect bacterial reduction. Results indicate that in the environments tested, the majority of the reduced iron found results from indirect reduction mediated by hydrogen sulfide, a result of dissolution and precipitation phenomena, or is a result of direct bacterial reduction using hydrogen as an electron donor. Direct iron reduction plays a minor role in carbon turnover in these environments.     

Shewanella oneidensis MR-1 uses overlapping pathways for iron reduction at a distance and by direct contact under conditions relevant for Biofilms

We developed a new method to measure iron reduction at a distance based on depositing Fe(III) (hydr)oxide within nanoporous glass beads. In this "Fe-bead" system, Shewanella oneidensis reduces at least 86.5% of the iron in the absence of direct contact. Biofilm formation accompanies Fe-bead reduction and is observable both macro- and microscopically. Fe-bead reduction is catalyzed by live cells adapted to anaerobic conditions, and maximal reduction rates require sustained protein synthesis. The amount of reactive ferric iron in the Fe-bead system is available in excess such that the rate of Fe-bead reduction is directly proportional to cell density; i.e., it is diffusion limited. Addition of either lysates prepared from anaerobic cells or exogenous electron shuttles stimulates Fe-bead reduction by S. oneidensis, but iron chelators or additional Fe(II) do not. Neither dissolved Fe(III) nor electron shuttling activity was detected in culture supernatants, implying that the mediator is retained within the biofilm matrix. Strains with mutations in omcB or mtrB show about 50% of the wild-type levels of reduction, while a cymA mutant shows less than 20% of the wild-type levels of reduction and a menF mutant shows insignificant reduction. The Fe-bead reduction defect of the menF mutant can be restored by addition of menaquinone, but menaquinone itself cannot stimulate Fe-bead reduction. Because the menF gene encodes the first committed step of menaquinone biosynthesis, no intermediates of the menaquinone biosynthetic pathway are used as diffusible mediators by this organism to promote iron reduction at a distance. CymA and menaquinone are required for both direct and indirect mineral reduction, whereas MtrB and OmcB contribute to but are not absolutely required for iron reduction at a distance.     

根系活动对湿地植物根际铁异化还原的影响及机制研究进展

根系活动是影响湿地植物根际铁异化还原速率的关键因素之一。以往国内外湿地铁异化还原的研究多为分析和比较各类中宏观生境中铁异化还原能力的差异。近年来,湿地植物根际微域铁的生物地球化学行为也日益成为该领域的研究热点。综述了根际铁异化还原研究概况,梳理了根系活动对根际铁异化还原关键因子的作用机制,分析了根际铁异化还原和其他有机质代谢途径的竞争关系,探讨了根际铁异化还原对根系活动动态变化和异质性的响应,提出了根际铁异化还原的概念模型,并指出了未来我国湿地植物根际铁异化还原研究应加强的工作。     

闭合复位与切开复位治疗桡骨远端骨折的疗效对比

目的:探讨闭合复位与切开复位对桡骨远端骨折患者的临床疗效。方法:回顾性分析2013年6月至2014年3月我院收治的60例桡骨远端骨折患者,随机数字表法分为切开复位组和闭合复位组,每组30例。闭合复位组患者给予闭合复位小夹板或石膏固定治疗,切开复位组患者给予切开复位内固定术治疗。观察并比较两组患者治疗前后掌倾角、尺偏角以及桡骨长度、术中出血量和手术时间,骨折愈合时间以及患者临床疗效进行检测并比较。结果:与治疗前相比,治疗后患者的掌倾角、尺偏角、桡骨长度水平均升高(P0.05);与闭合复位组相比,切开复位组患者掌倾角、尺偏角、桡骨长度评分水平较高(P0.05),术中出血量、手术时间水平较高(P0.05),临床治疗的优良率较高(P0.05),两组患者的骨折愈合时间无显著差异(P0.05)。结论:与闭合复位相比,切开复位能够明显恢复桡骨远端骨折患者的掌倾角、尺偏角以及桡骨长度,但手术时间以及术中出血量较多,临床疗效较好,两组患者的骨折愈合时间无明显差异。     

Increased nitroblue tetrazolium dye reduction by human neutrophils stimulated by interferon in vitro

Human leukocyte interferon enhanced nitroblue tetrazolium dye (NBT) reduction by human neutrophils (PMNs). Increase in NBT reduction paralleled increase in interferon dose. When human leukocyte interferon was heated to 60 C or 80 C for 30 min, both the antiviral activity and the effect on NBT reduction decreased. Human leukocyte interferon neutralized with anti-human leukocyte interferon serum showed no effect on NBT reduction. A human fibroblast interferon preparation also enhanced NBT reduction. The species dependency of interferon was shown in NBT reduction as well as in antiviral activity.     

Silicomolybdate reduction by isolated pea chloroplasts

Conditions for the optimization of silicomolybdate reduction by isolated pea chloroplasts are described. Maximum rates of reduction are related to time of addition to the chloroplasts and the presence of an oxidizing cofactor, such as ferricyanide. Silicomolybdate or silicomolybdate plus ferricyanide reduction is only partially inhibited by a concentration of CMU which totally abolishes ferricyanide reduction. Evidence for a differing response of the two reduction sites to silicomolbydate is described.     

Selenium reduction by a denitrifying consortium

A denitrifying bacterial consortium obtained from the Pullman, Washington wastewater treatment facility was enriched under denitrifying conditions and its ability to reduce selenite and selenate was studied. Replicate experiments at two different experimental conditions were performed. All experiments were performed under electron-acceptor limiting conditions, with acetate as the carbon source and nitrate the electron acceptor. In the first set of experiments, selenite was present, whereas, in the second set, selenate was added. A significant lag period of approximately 150 h was necessary before selenite or selenate reduction was observed. During this lag period, nitrate and nitrite use was observed. Once selenite or selenate reduction had started, nitrate and nitrite reduction was concomitant with selenium species reduction. Trace amounts of selenite were detected during the selenate reduction study. Analysis of the data indicates that, once selenium species reduction was induced, the rate of reduction was proportional to the selenium species concentration and to the biomass concentration. Furthermore, at similar biomass and contaminant concentrations, selenite reduction is approximately four times faster than selenate reduction. Copyright 1999 John Wiley & Sons, Inc.     

Competitive Mechanisms for Inhibition of Sulfate Reduction and Methane Production in the Zone of Ferric Iron Reduction in Sediments

Mechanisms for inhibition of sulfate reduction and methane production in the zone of Fe(III) reduction in sediments were investigated. Addition of amorphic iron(III) oxyhydroxide to sediments in which sulfate reduction was the predominant terminal electron-accepting process inhibited sulfate reduction 86 to 100%. The decrease in electron flow to sulfate reduction was accompanied by a corresponding increase in electron flow to Fe(III) reduction. In a similar manner, Fe(III) additions also inhibited methane production in sulfate-depleted sediments. The inhibition of sulfate reduction and methane production was the result of substrate limitation, because the sediments retained the potential for sulfate reduction and methane production in the presence of excess hydrogen and acetate. Sediments in which Fe(III) reduction was the predominant terminal electron-accepting process had much lower concentrations of hydrogen and acetate than sediments in which sulfate reduction or methane production was the predominant terminal process. The low concentrations of hydrogen and acetate in the Fe(III)-reducing sediments were the result of metabolism by Fe(III)-reducing organisms of hydrogen and acetate at concentrations lower than sulfate reducers or methanogens could metabolize them. The results indicate that when Fe(III) is in a form that Fe(III)-reducing organisms can readily reduce, Fe(III)-reducing organisms can inhibit sulfate reduction and methane production by outcompeting sulfate reducers and methanogens for electron donors.     

Chemical Characterisation of Organic Electron Donors For Sulfate Reduction For Potential Use in Acid Mine Drainage Treatment

The production of acid mine drainage (AMD) containing high amounts of sulfate, heavy metals and low pH is of increasing concern. AMD is highly corrosive and results in economic and environmental problems. Organic electron donors for sulfate reduction were chemically characterised for potential use in AMD treatment. This was done in a process to develop a correlation between chemical composition and the capacity to drive sulfate reduction. Potential organic electron donors for sulfate reduction were chemically characterised in terms of dry matter content, ash content, total Kjeldahl nitrogen, lignin content, cellulose content, crude fat, crude fibre, in vitro digestibility, water-soluble carbohydrates, total non-structural carbohydrates and starch content. The chemical composition of the organic electron donors was then compared to results obtained from pilot plant studies where the organic electron donors for sulfate reduction were evaluated in terms of sulfate reduction. The chemical composition of the carbon source severely impacted its capacity to drive sulfate reduction and may be used to assist in predicting the sulfate reduction capacity of a carbon source. Organic electron donors for sulfate reduction high in protein content and low in lignin content or high in carbohydrate and crude fat content increased the capacity of a carbon source to drive sulfate reduction. The higher the fibre content of a carbon source, the lower the capacity to drive sulfate reduction. No correlation could be drawn between % dry matter, % ash content and sulfate reduction for the organic electron donors tested. Chemical characterisation can be used to assist in predicting sulfate reduction capacity of organic electron donors.     

Methemoglobin reduction under near physiological conditions

Pure methemoglobin was prepared from fresh red cells and was used as substrate for methemoglobin reduction reaction. Two sources of methemoglobin reductase were used: (a) red cell hemolysate which was prepared by freezing and thawing of unwashed red cells; (b) purified methemoglobin reductase from bank blood. Methemoglobin reduction rate was measured in a mixture of pure methemoglobin (substrate) and hemolysate (enzyme). In other experiments the rate of methemoglobin reduction was measured in the above mixture with the addition of various other compounds such as NADH, cytochrome b5, and pure methemoglobin reductase. Only the addition of pure enzyme accelerated the rate of methemoglobin reduction. In other experiments, the rate of methemoglobin reduction was measured when the reduction reaction was carried out in the presence of various amounts of deoxyhemoglobin, globin, or albumin. It was shown that all proteins tested here decreased the reduction rate. It is concluded that (a) in the red cell, under normal conditions, only the activity of the methemoglobin reductase controls the speed of methemoglobin reduction, and (b) the inhibition of methemoglobin reduction by reduced hemoglobin is mostly nonspecific suggesting a noncompetitive reaction.