Heme P460 of Hydroxylamine Oxidoreductase of Nitrosomonas

Hooper, Alan B.; Debey, Pascale; Andersson, Karin; Balny, Claude

doi:10.1111/j.1432-1033.1983.tb07534.x

Cited by 29 publications

(23 citation statements)

References 22 publications

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“…The slope of the latter curves gave second-order rate constants kl,co. At atmospheric pressure and for a given temperature, the values of both k,,,, and AH* (Table 1) were close to published values [6]. The twofold difference observed may be due to the fact that the present experiments were carried out in Tris rather than phosphate buffer.…”

Section: Kinetics Of Formation Of the Co-complex With Ferrous P460supporting

confidence: 67%

“…The presence of organic co-solvent (40% ethylene glycol) increased kl,co by a factor of 2.5-3. Similar solvent effects have been observed in this enzyme [6] and many other enzymes [13].…”

Section: Kinetics Of Formation Of the Co-complex With Ferrous P460supporting

confidence: 57%

“…Mixing of dithionite-reduced enzyme with CO solution resulted in a progressive increase in A444 and a concomitant decrease in A465 as CO bound to P460 [6]. The kinetics were studied in either buffered aqueous solution, pH 7.5, or 40% (by vol.)…”

Section: Kinetics Of Formation Of the Co-complex With Ferrous P460mentioning

confidence: 99%

“…Values of association rate constant kl,co of the binding of CO with the reduced enzyme were determined as described previously [6]. The kinetics of reduction of c hemes of the enzyme with NHzOH consisted of two phases.…”

Section: Determination Of Kinetic Constants and Activation Volumementioning

confidence: 99%

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Effect of solvent, pressure and temperature on reaction rates of the multiheme hydroxylamine oxidoreductase

Balny

Hooper

1988

European Journal of Biochemistry

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Hydroxylamine oxidoreductase (HAO) of the ammonia-oxidizing bacterium Nitrosomonas catalyzes the oxidation: N H 2 0 H + H 2 0 + HNOz + 2e-+ 2 H'. The heme-like chromophore P460 is part of a site which binds substrate, extracts electrons and then passes them to the many c hemes of the enzyme. Reduction of the c hemes by hydroxylamine is biphasic with apparent first-order rate constants kl and k z . CO binds to ferrous P460 with apparent first-order rate constants, kl,co. In this work we have measured the binding of CO to ferrous P460 of hydroxylamine oxidoreductase and the reduction by substrate of some of the 24 c hemes of the ferric enzyme. These reactions have been studied in water and 40% ethylene glycol, at temperatures ranging from -15°C to 2 0 . 7 T and at hydrostatic pressures ranging over 0.1 -80 MPa. From the measurements, thermodynamic parameters A V* (activation volume), AG*, AH', and AS* have been calculated.CO binding. Binding of CO to ferrous P460 was similar to the binding of CO to ferrous horseradish peroxidase. The change of solvent had only a limited effect on A V* (-30 ml . mol-I), AG", AH* or AS* and did not cause an inflection in the Arrhenius plot or downward displacement of the linear relationship between In kl,co and P at a critical temperature. Binding was exothermic at high temperatures. The response of the binding of CO to solvent, temperature and pressure suggested that the CO binding site had little access to solvent and was not susceptible to change in protein conformation.Fast phase of reduelion of c hemes. Changing the solvent from water to 40% ethylene glycol resulted in a decrease from 90% to 50% in the relative number of c hemes reduced during the fast phase, an increase in activation volume from -3.6 ml . mol-' to 57 ml . mol-' and changes in other thermodynamic parameters. The activation volume increased with decreasing temperature. The Arrhenius plot had a downward inflection at about 0°C and, in water or ethylene glycol, the linear dependence of In kl on P was displaced downwards as the temperature changed from 3.5"C to -15°C.Slow phase of reduction of c hemes. Changing the solvent from water to 40% ethylene glycol resulted in an increase in the relative number of c hemes reduced during the slow phase from 10% to 50%. The activation volume, which was not measurable in water because of the low absorbance change, was -30 ml . mo1-l in ethylene glycol. The activation volume increased with increasing temperature. The Arrhenius plot had a downward inflection at about 0 "C yet the linear dependence of In k2 on P had no downward displacement in this temperature range. At high pressure and low temperatures the rate of heme reduction increased with decreasing temperature. In contrast to the reaction of CO with the substrate site, the results suggested that changes in enzyme conformation imposed by changes in solvent, temperature or pressure affected the rates of intramolecular electron transfer from the substrate site to c hemes.Hydroxylamine oxidoreductase from the ammonia-oxidizin...

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Section: Kinetics Of Formation Of the Co-complex With Ferrous P460supporting

confidence: 67%

“…The presence of organic co-solvent (40% ethylene glycol) increased kl,co by a factor of 2.5-3. Similar solvent effects have been observed in this enzyme [6] and many other enzymes [13].…”

Section: Kinetics Of Formation Of the Co-complex With Ferrous P460supporting

confidence: 57%

Section: Kinetics Of Formation Of the Co-complex With Ferrous P460mentioning

confidence: 99%

Section: Determination Of Kinetic Constants and Activation Volumementioning

confidence: 99%

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Effect of solvent, pressure and temperature on reaction rates of the multiheme hydroxylamine oxidoreductase

Balny

Hooper

1988

European Journal of Biochemistry

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“…Kinetics of reduction and reoxidation Kinetic measurements and rapid-scan absorption spectra were made with a thermostated anaerobic stopped-flow mixing apparatus [15] adapted to the Union Giken model RA 41 5 -RA 401 (Osaka, Japan) fast-response spectrophotometer as described previously [16,17]. Kinetic constants (standard deviation 10%) were calculated with the aid of a SORD (Japan) microcomputer on the basis of a fitting program developed by Union Giken.…”

Section: Materials and Methods Enzymes And Chemicalsmentioning

confidence: 99%

Conformational change accompanies redox reactions of the tetraheme cytochrome c-554 of Nitrosomonas europaea

DiSpirito¹,

Balny²,

Hooper³

1987

Eur J Biochem

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Cytochrome c-554 of the ammonia-oxidizing chemolithoautotropic bacteria is thought to mediate electron transfer from hydroxylamine oxidoreductase to a terminal oxidase and/or to ammonia monooxygenase. The cytochrome has four c hemes which interact magnetically and have the same redox potential.We report that the kinetics of reduction of ferric cytochrome c-554 by dithionite or the oxidation of ferrous cytochrome c-554 by O2 or H 2 0 2 are complex and multiphasic. Transient rapid-scan difference spectra indicate discrete maxima at approximately 418 nm, 425 nm and 432 nm. Absorbance changes at all three difference maxima appear to occur in all kinetic phases, although not in equal amounts for each wavelength.Reduction by 20 mM dithionite was biphasic. At pH 7.5 the first phase, which involved approximately 50% of the total absorbance change, had a rate constant (20°C) of 140 s -l and energy of activation of 20 kJ . mol-'.The slow phase had a rate constant 0.43 sC1 and a relatively high energy of activation, 87 kJ . mol-l, suggesting that a change in protein configuration accompanied the reaction. As the pH of the solution increased, the rate constant for both phases decreased and the fraction of absorbance change in the rapid phase increased. Oxidation of ferrous cytochrome c-554 by O2 involved a discrete rapid phase with a rate constant of 14 s -l , accounting for 6% of the absorbance. The remainder of the reaction was multiphasic with rate constants in the range 0.1 -0.01 s-'. With H 2 0 2 as the oxidant, the rapid phase involved 39% of the change in absorbance with a rate constant of 19 s-'. The remainder of the reoxidation was multiphasic with rate constants ranging over 0.4-0.01 s-'.Oxidation of NH3 to HN02 by Nitrosomonas europaea appears to involve the sequential action of a membrane ammonia monooxygenase (AMO) (NH3 + 2e-+ 2H' + O2 + N H 2 0 H + H20, [I]) and hydroxylamine oxidoreductase (HAO) (NH20H + H 2 0 + HNOz + 4e-+ 4H+, [2]). The latter enzyme is a complex multiheme protein [3] which is reduced by NH20H and is presumed to donate two electrons to the ammonia monoxygenase and two electrons to a membrane terminal oxidase (2H' + 2e-+ 1/2 O2 -, H 2 0 [4]).

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Hydroxylamine Oxidoreductase

Igarashi¹,

Tanaka²

2004

Handbook of Metalloproteins

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as hydroxylamine oxidase (HAO).Hydroxylamine oxidoreductase (HAO) catalyzes the fourelectron oxidation of hydroxylamine to nitrite. The enzyme shows a high biologically specificity. The most efficient oxidizing substrate is hydroxylamine, and no other nitrogen compound except for hydrazine reacts with 3D Structure Schematic representation of the trimer structure of HAO from N. europaea. Each subunit is a single color (yellow, blue, pink). The hemes are shown in red ball-and-stick representation. Prepared with programs molscript 58 and raster3d. 59 The PDB code is 1FGJ.

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Heme P460 of Hydroxylamine Oxidoreductase of Nitrosomonas

Cited by 29 publications

References 22 publications

Effect of solvent, pressure and temperature on reaction rates of the multiheme hydroxylamine oxidoreductase

Effect of solvent, pressure and temperature on reaction rates of the multiheme hydroxylamine oxidoreductase

Conformational change accompanies redox reactions of the tetraheme cytochrome c-554 of Nitrosomonas europaea

Hydroxylamine Oxidoreductase

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