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Y Orii

Publications and source records attributed to Y Orii.

At least 37 records · Page 2Linked to original sources
PubMed1994

Facilitated intramolecular electron transfer in cytochrome bo-type ubiquinol oxidase initiated upon reaction of the fully reduced enzyme with dioxygen.

Flow-flash and double-flash studies of the reaction of fully reduced bo-type quinol oxidase with oxygen have revealed that a single turnover of the enzyme proceeds much faster than mammalian cytochrome c oxidase. Facilitated intramolecular electron transfer in the bo-type oxidase with k > 5 x 10(4) s-1 at pH 7.4 and 20 degrees C is responsible for this fast turnover. The kinetics of this reaction indicates that the oxygen reduction does not require electron exchange between quinol oxidase molecules, each having three metal centers. Thus, a bound quinol in the fully reduced enzyme is suggested to be an electron source for complete reduction of dioxygen into water supplementing electrons provided by the metal centers. A single turnover of the quinol oxidase yields a novel spectral species with a Soret maximum at 415 nm corresponding to a 'pulsed' state of mammalian cytochrome c oxidase.

Animals↗
PubMed1994

Reactions of the Escherichia coli flavohaemoglobin (Hmp) with oxygen and reduced nicotinamide adenine dinucleotide: evidence for oxygen switching of flavin oxidoreduction and a mechanism for oxygen sensing.

The soluble flavohaemoglobin (Hmp) of Escherichia coli contains haem B and FAD in a single 44 kDa polypeptide, and shows NADH oxidase activity. The oxidized protein reacted rapidly with NADH in the presence of O2 to form an oxygenated species while the flavin remained largely oxidized. Spectral and kinetic analyses revealed rapid biphasic reduction and oxygenation of high-spin haem with apparent relaxation times of 6 and 64 ms at pH 8 and 25 degrees c, suggestive of a significant physiological role for the protein. This was followed by a monophasic reduction of the flavin with a relaxation time of 92 ms. On exhaustion of oxygen, the oxygenated haem was converted into the deoxy form biphasically with relaxation times of 43 and 170 s, followed by extensive reduction of the flavin with corresponding relaxation times of 70 and 256 s. Based on these observations, we propose that Hmp could act as an oxygen sensor in E. coli by combining with intracellular oxygen, thus limiting flavin reduction in the aerobic steady state. Lowering of the oxygen concentration causes dissociation of the oxy species and sustained flavin reduction. Because Hmp can reduce Fe(III), such a mechanism might control, for example, flavin-mediated Fe(III) reduction required for activation of the anaerobic gene regulator, Fnr.

Bacterial Proteins↗
PubMed1994

Membrane potential-linked reversed electron transfer in the beef heart cytochrome bc1 complex reconstituted into potassium-loaded phospholipid vesicles.

The cytochrome bc1 complex purified from beef heart mitochondria was incorporated into potassium (K+)-loaded phospholipid vesicles by a cholate dialysis method to study the reverse reaction of electron transfer in the complex. The reduction of cytochrome b in the presence of sodium ascorbate was observed on addition of valinomycin to the K(+)-loaded proteoliposomes in a medium containing no external KCl; it was followed by the gradual oxidation. Nigericin accelerated the reoxidation of reduced cytochrome b, indicating that a K+ diffusion potential (negative inside) induced the reduction of cytochrome b. The extent of the cytochrome b reduction depended on the magnitude of the diffusion potential across the liposomal membranes, and its maximal reduction was attained at more than 210 mV of the diffusion potential. It was cytochrome b562 that was reduced during the establishment of the K+ diffusion potential in the presence of ascorbate, and about 90% of cytochrome b562 was estimated to be reduced. Antimycin A and myxothiazol inhibited the diffusion potential-induced reduction of cytochrome b562, and ubiquinone was proved to be essential for the reversed electron transfer. The K+ diffusion potential also induced the partial reduction of cytochrome b566 when cytochrome b562 had previously been reduced with ascorbate plus tetramethyl-p-phenylenediamine. These results were interpreted well based on the Q cycle scheme which assumed the energy-dependent reduction of ubiquinone at center o. Dicyclohexylcarbodiimide, which did not perturb the ability of proteoliposomes to generate the K+ diffusion potential, inhibited the energy-dependent reduction of cytochrome b562 without a significant loss in the catalytic activity of the complex. The half-inhibition was brought about by 200 mol of dicyclohexylcarbodiimide/mol of cytochrome c1. These results strongly suggest the coupling of a proton flow with the reversed electron transfer in the bc1 complex.

Animals↗
PubMed1993

Immediate reduction of cytochrome c by photoexcited NADH: reaction mechanism as revealed by flow-flash and rapid-scan studies.

Upon exposure of an aqueous solution of NADH and cytochrome c to a laser pulse at 355 nm under anaerobic conditions, cytochrome c is reduced within 1-2 ms to a maximal extent of 90%. The reduction proceeds in two phases: rapid reduction by hydrated electrons followed by bimolecular electron transfer from the NAD radical to ferric cytochrome c. In addition, a third reduction phase emerges in the presence of an appropriate concentration of molecular oxygen, where the superoxide anion is a reductant. As the oxygen concentration approaches 20% saturation, the cytochrome c reduction by NAD is abolished first and then the reduction by hydrated electrons, since molecular oxygen competes with cytochrome c for NAD and hydrated electrons. At 20% oxygen, cytochrome c is reduced almost exclusively by the superoxide anion, but the amount reduced on a single laser pulse is only one-fourth that reduced under anaerobic conditions. The second-order rate constants for the reduction of cytochrome c at pH 7.4 and 20 degrees C by NAD and the superoxide anion are 2.0 x 10(9) and 4.0 x 10(6) M-1 s-1, respectively.

Animals↗
PubMed1993

Superoxide-producing cytochrome b. Enzymatic and electron paramagnetic resonance properties of cytochrome b558 purified from neutrophils.

Molecular properties of superoxide (O2-)-producing cytochrome b558 purified from neutrophils were investigated focusing on the mechanism of the catalytic reaction. The purified cytochrome, which was depleted of FAD, exhibited high O2(-)-generating activity with consumption of NADPH in the presence of microsomal NADPH-cytochrome P-450 reductase. Exogenous additions of CO, CN-, or N3- had no effect on the enzymatic activity. Potentiometric titration of the ferric-ferrous couple of the cytochrome showed that the midpoint reduction potential was -255 mV at pH 7.4. When the reaction of the reduced cytochrome with O2 was analyzed by stopped flow and rapid scanning spectrophotometry, the ferrous form was found to be converted to the ferric form at a rate constant of 9.3 x 10(6) M-1 s-1 at 10 degrees C without showing formation of an oxygenated intermediate. EPR measurement of the ferric cytochrome at 10 K showed that the electronic spin state was in a low spin with g values of 3.2, 2.05, and 1.5. These results suggest that the heme in a six-coordinated low spin state catalyzes one electron reduction of O2 without ligation of O2 to the heme iron during the catalytic cycle.

Animals↗
PubMed1992

The oxygenated flavohaemoglobin from Escherichia coli: evidence from photodissociation and rapid-scan studies for two kinetic and spectral forms.

The kinetics of dissociation and reassociation of the oxygenated species of Escherichia coli flavohaemoglobin (Hmp) were studied using stopped-flow rapid-scan and flash photolysis spectrophotometry at 25 degrees C. The oxygenated compound(s) form rapidly on mixing oxygen with the NADH-reduced flavohaemoglobin. On exhaustion of NADH, with residual oxygen, decay occurs in two phases to give a form in which haem b and flavin are oxidized. Spectral changes during this process suggest a direct release of O2- from the oxy form. Photodissociation of the oxygenated species generates the unliganded protein, which recombines with oxygen to give two spectrally and kinetically distinct forms. The reversibility of the oxygen reaction and the rapid reassociation kinetics after photodissociation confirm the haemoglobin-like features of this protein.

Bacterial Proteins↗
PubMed1992

Acceleration of the oxygen reaction in CuA-deficient Nitrosomonas europaea cytochrome c oxidase as revealed by the flow-flash measurement.

The oxygen reaction of Nitrosomonas europaea cytochrome c oxidase containing either 2Cu or 1Cu per two heme a molecules was investigated by the flow-flash technique at 20 degrees C. The reaction profiles of the bacterial enzyme were essentially the same as those of bovine heart cytochrome c oxidase, although the rate of the primary oxygen compound formation was much slower. The 1Cu enzyme exhibited higher rates for both primary oxygen compound formation and intramolecular electron transfer than the 2Cu enzyme. This result clearly indicates that CuA is not essential functionally for the oxidation of ferrous heme a moieties, and suggests its structural importance in maintaining the molecular integrity of N. europaea cytochrome oxidase.

Copper↗
PubMed1991

Stopped-flow and rapid-scan studies of the redox behavior of cytochrome aco from facultative alkalophilic Bacillus.

Cytochrome aco purified from an alkalophilic bacterium grown at pH 10 contains hemes a, b, and c as prosthetic groups, and their redox behavior was examined by using stopped-flow and rapid-scan techniques. Under anaerobic conditions the reduction of both heme a and c moieties with dithionite proceeded exponentially but with different rates, usually the former being reduced about 4 times faster than the latter. The reduction of protoheme was much slower, and a time-difference spectrum for this species was of a high spin type with absorption peaks at 433, 557, and 609 nm. Only the protoheme combined with CO, fulfilling the criteria for cytochrome o. Potentiometric titrations determined a midpoint potential of c heme to be 95 mV at pH 7.0 and 25 degrees C and suggested the presence of two forms of a heme with midpoint potentials of 250 and 323 mV. Cytochrome aco utilizes ascorbate plus N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD) to reduce oxygen relatively rapidly without added cytochrome c (Qureshi, M. H., Yumoto, I., Fujiwara, T., Fukumori, Y., Yamanaka, T. (1990) J. Biochem. 107, 480-485). During the steady state, however, heme a stayed almost fully reduced in contrast to a partial reduction of heme c. Even after exhaustion of the dissolved oxygen the extent of reduction of heme c was 60-70% that attained by the dithionite reduction. When ascorbate plus TMPD-reduced cytochrome aco was exposed to oxygen the reduced heme c was oxidized rapidly whereas the oxidation of reduced a heme was negligibly slow. The full reduction of heme a during the steady state and its extremely slow oxidation rendered participation of heme a in the oxidase reaction less likely. A novel peak appearing transiently around 567 nm during the reaction was tentatively ascribed to an intermediate form of protoheme, or o heme, which was thus supposed to react directly with molecular oxygen. These results suggest strongly that the main electron transfer pathway would be c----o----oxygen. A possible role of a in regulating the electron flow through the main pathway and its functional relationship to a heme in the aa3-type cytochrome oxidase were discussed.

Ascorbic Acid↗
PubMed1990

Formation and decay of cytochrome c peroxidase compound ES during aerobic reduction with dithionite.

Stopped-flow and rapid scanning studies have clearly demonstrated that mixing of an oxygen-saturated solution of yeast cytochrome c peroxidase with sodium dithionite yields compound ES, indicating generation of H2O2. The formation of compound ES was most pronounced when [Na2S2O4]/[O2] approximately 1, and it reverted to the ferric form while standing. Even in the presence of an excess of dithionite ([Na2S2O4]/[O2] = 3.4) compound ES was formed immediately, but was soon replaced by the ferric form, followed by its final reduction to the ferrous state. The apparent first order rate constant for the decay of compound ES to the ferric form increased linearly with the square root of the dithionite concentration, thus involvement of SO2- in that process being suggested.

Aerobiosis↗
PubMed1990

Cytochrome c oxidase activity in human liver specimens. An index of prognosis for hepatic resection.

Cytochrome c oxidase activity, measured as Vmax (nanomoles of cytochrome c oxidized per second per milligram of protein) and Michaelis constant (Km) (micromoles per liter), was measured spectrophotometrically in human liver specimens obtained by needle biopsy from 43 patients. In 16 normal livers, the Vmax and Km values ranged from 1.26 to 2.25 nmol/s per milligram of protein and from 2.78 to 3.95 mumol/L, respectively. In 27 patients with liver cirrhosis or chronic hepatitis, these values ranged from 1.60 to 3.80 nmol/s per milligram of protein and from 2.80 to 6.50 mumol/L, respectively. Patients with Vmax above 2.5 nmol/s per milligram of protein or Km above 5.0 mumol/L had a high incidence of postoperative complications even after minor hepatic resection. By contrast, even patients with liver cirrhosis or chronic hepatitis could tolerate major hepatic resection as long as their Vmax and Km values were within the normal range. These findings indicate that the cytochrome c oxidase activity in liver specimens can serve as a prognostic sign in hepatic resection even in patients with liver cirrhosis or chronic hepatitis.

Adult↗
PubMed1990

Specific inhibition of redox-linked proton pump activity of cytochrome oxidase by oleate hydroperoxide and involvement of ferrocytochrome c in the catabolism of hydroperoxide.

Both oleic acid and oleate hydroperoxide at concentrations below 200 nmol/mg asolectin remarkably depressed the proton pumping of cytochrome c oxidase reconstituted into liposomes but did not affect the respiratory control ratio. The inhibitory effect was comparable to that of N,N'-dicyclohexylcarbodiimide. Oleate hydroperoxide in the vesicles was reduced by ferrocytochrome c in the absence of cytochrome oxidase and converted to the hydroxy fatty acid. This non-enzymatic oxidation of ferrocytochrome c affected slightly the proton pumping and the cytochrome c oxidation by liposomal cytochrome oxidase. A physiological role of ferrocytochrome c in catabolism of the hydroperoxide of fatty acids is thus suggested.

Animals↗
PubMed1989

Effect of the distal histidine modification (Cyanation) of myoglobin on the ligand binding kinetics and the heme environmental structures.

The kinetics of carbon monoxide (CO) binding to myoglobin (Mb) modified at the distal histidine (His) by cyanogen bromide (BrCN) has been studied. The CO association and dissociation rates of BrCN-modified Mb were obtained as 1.8 x 10(3) M-1 s-1 and 0.13 s-1, respectively (20 degrees C and pH 7.0). Thermodynamic parameters were obtained as well. These values are notable, compared with those for other hemoproteins, the slowest association and the fastest dissociation rates among various hemoproteins examined so far. On the basis of the available structural data obtained from the absorption, 1H NMR, and IR spectral measurements, these unique kinetic and thermodynamic properties were reasonably explained in terms of the steric restriction at the modified distal side.

Animals↗
PubMed1989

Ubiquitous formation of catalase compound II in hemoglobin-free perfused rat liver and detection of novel spectral species.

Spectral examinations of hemoglobin-free perfused rat livers with a high-sensitivity reflectance spectrophotometer have revealed an accumulation of catalase Compound II to an amount comparable to that of Compound I under the aerobic steady state. This finding is in contrast to a recent proposal that NADPH associated with catalase both prevents and reverses the accumulation of Compound II (Kirkman, H. N., Galiano, S., and Gaetani, G.F. (1987) J. Biol. Chem. 262, 660-666). Furthermore, spectral species with a broad peak extending from 550 to 600 nm were observed in a time range between the methanol-induced decays of Compound I and Compound II. When rats were treated with 3-amino-1,2,4-triazole, catalase Compound I was not detected but spectral species with peaks at 570, 556 and 530 nm were observed. These novel spectral profiles suggest contributions from "peroxy" and "ferryl" forms of cytochrome oxidase.

Amitrole↗
PubMed1989

Effects of hypoxia on noradrenaline release and neuronal reuptake in isolated rabbit thoracic aortic strips.

To clarify the effects of hypoxia on stimulus-evoked noradrenaline release and on neuronal reuptake of the released noradrenaline, we examined the effects of hypoxia on contraction responses of rabbit thoracic aortic strips to transmural electrical stimulation and on the stimulation-evoked overflow of total [3H] and [3H]noradrenaline from the strips prelabelled with [3H]noradrenaline. This was done in the presence or absence of an inhibitor of neuronal uptake (cocaine). In a medium equilibrated with a gas mixture of 95% O2/5% CO2 (control), cocaine doubled the stimulation-evoked overflow of total [3H] and [3H]noradrenaline; there was a concomitant increase (130%) in contractions to electrical stimulation. At 0% O2 (95% N2/5% CO2, hypoxia), cocaine had no significant effects on either the stimulation-evoked overflow of total [3H] and [3H]noradrenaline or contractions. In the absence of the drug, hypoxia decreased the stimulation-evoked overflow of total [3H] and [3H]noradrenaline to 47% and 43%, respectively, of the control values, whereas these values were 31% and 28%, respectively, after exposure to cocaine. The inhibition by hypoxia of contraction responses to electrical stimulation was greater in the presence of cocaine than in its absence. These results show that hypoxia inhibits both noradrenaline release evoked by a given stimulus and neuronal uptake.

Animals↗
PubMed1988

Determination of cytochromes in human liver contaminated with hemoglobin.

A spectrophotometric method was developed to determine cytochromes in human liver mitochondria contaminated with hemoglobin. The influence of hemoglobin on the measurement was canceled by keeping hemoglobin in the carbon monoxide bound form throughout determination. Mitochondria were solubilized by 2% sodium cholate, and cytochromes were reduced enzymatically with glutamate and succinate as substrates to the maximal extent. The amount of cytochromes determined spectrophotometrically was linearly correlated with mitochondrial protein at least up to the extent of 8 mg/ml.

Aged↗
PubMed1988

Effects of hypoxia on contractile responses of rabbit aortic strips to transmural electrical stimulation.

To clarify the effects of hypoxia on adrenergic transmission, we examined the contractile responses of isolated rabbit aortic strips to electrical stimulation, the concentration-response relationships for noradrenaline and KCl, and the electrical stimulation-evoked overflows of total [3H] and [3H]noradrenaline from strips preloaded with [3H]noradrenaline in media equilibrated with gas mixtures containing various concentrations of O2. Contractile responses to electrical stimulation were completely inhibited by tetrodotoxin and alpha-adrenoceptor antagonists such as phentolamine and phenoxybenzamine, but were not affected by indomethacin. When the concentration of O2 in the gas mixture was decreased from 95% to 20%, the contractile responses to electrical stimulation remained unchanged, but as the concentration of O2 was further decreased, the responses were inhibited concentration-dependently. At 0% O2, the response was inhibited by about 80% when compared with control values obtained at 95% O2, and the electrical stimulation-evoked overflows of total [3H] and [3H]noradrenaline into the superfusates were decreased by about 55%. At 0% O2, the concentration-response curve for exogenous noradrenaline was shifted to the right about 50-fold and the maximum response was decreased by 25%. The maximum contractile responses of aortic strips from animals pretreated with reserpine or 6-hydroxydopamine to high KCl were decreased slightly (about 15%). These results suggest that inhibition of adrenergic transmission under hypoxic conditions is mainly the result of a decrease in the stimulus-evoked release of noradrenaline and of a decrease in the affinity of alpha-adrenoceptor for noradrenaline and/or inhibition of signal transduction mechanisms, although hypoxia also causes a slight decrease in the contractility of vascular smooth muscle.

Animals↗