scholarly journals Metal-binding mechanism of Cox17, a copper chaperone for cytochrome c oxidase

2004 ◽  
Vol 382 (1) ◽  
pp. 307-314 ◽  
Author(s):  
Peep PALUMAA ◽  
Liina KANGUR ◽  
Anastassia VORONOVA ◽  
Rannar SILLARD
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Metal Binding ◽  
Copper Ions ◽  
Copper Chaperone ◽  
Copper Chaperones ◽  
Rate Limiting Step ◽  
Charge State Distributions ◽  
Rate Limiting ◽  
Partner Proteins

Cox17, a copper chaperone for cytochrome c oxidase, is an essential and highly conserved protein. The structure and mechanism of functioning of Cox17 are unknown, and even its metalbinding stoichiometry is elusive. In the present study, we demonstrate, using electrospray ionization–MS, that porcine Cox17 binds co-operatively four Cu+ ions. Cu4Cox17 is stable at pH values above 3 and fluorescence spectra indicate the presence of a solvent-shielded multinuclear Cu(I) cluster. Combining our results with earlier EXAFS results on yeast CuCox17, we suggest that Cu4Cox17 contains a Cu4S6-type cluster. At supramillimolar concentrations, dithiothreitol extracts metals from Cu4Cox17, and an apparent copper dissociation constant KCu=13 fM was calculated from these results. Charge-state distributions of different Cox17 forms suggest that binding of the first Cu+ ion to Cox17 causes a conformational change from an open to a compact state, which may be the rate-limiting step in the formation of Cu4Cox17. Cox17 binds non-co-operatively two Zn2+ ions, but does not bind Ag+ ions, which highlights its extremely high metal-binding specificity. We further demonstrate that porcine Cox17 can also exist in partly oxidized (two disulphide bridges) and fully oxidized (three disulphide bridges) forms. Partly oxidized Cox17 can bind one Cu+ or Zn2+ ion, whereas fully oxidized Cox17 does not bind metals. The metal-binding properties of Cox17 imply that, in contrast with other copper chaperones, Cox17 is designed for the simultaneous transfer of up to four copper ions to partner proteins. Metals can be released from Cox17 by non-oxidative as well as oxidative mechanisms.

Is the Internal Electron Transfer the Rate-Limiting Step in the Catalytic Cycle of Cytochrome c Oxidase?

1988 ◽  
Vol 550 (1 Cytochrome Ox) ◽  
pp. 161-166 ◽  
Author(s):  
PAOLO SARTI ◽  
GIOVANNI ANTONINI ◽  
RANCESCO MALATESTA ◽  
BEATRICE VALLONE ◽  
MAURIZIO BRUNORI
Keyword(s):  
Electron Transfer ◽  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Catalytic Cycle ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Internal Electron ◽  
Rate Limiting

scholarly journals Transient kinetics of subunit-III-depleted cytochrome c oxidase

1986 ◽  
Vol 234 (3) ◽  
pp. 569-572 ◽  
Author(s):  
F Malatesta ◽  
G Antonini ◽  
P Sarti ◽  
M Brunori
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Flash Photolysis ◽  
Order Rate Constant ◽  
Kinetic Properties ◽  
Transient Kinetics ◽  
Rate Limiting Step ◽  
Subunit Iii ◽  
The Stability ◽  
Rate Limiting

Cytochrome c oxidase from ox heart was depleted of subunit III and its transient kinetic properties studied by stopped-flow and flash photolysis. It was found that the overall mechanism of electron transfer is very similar for subunit-III-depleted and native oxidase, although significant differences in some kinetic parameters have been detected. These include the second-order rate constant for cytochrome c oxidation and the rate-limiting step of the overall process. Moreover, at low cytochrome c/oxidase ratios (where the number of reducing equivalents is insufficient), the rate of reoxidation of cytochrome a was found to be very slow, even in air, and in fact for the subunit-III-depleted enzyme is even slower than for the native oxidase. The stability of reduced cytochrome a excludes the likelihood that removal of subunit III leads to a new O2-binding site, and the result may be relevant to the lowered vectorial H+/e- stoichiometry. The subunit-III-depleted oxidase can be pulsed under appropriate conditions and its combination with CO is unchanged, as shown by kinetic experiments and difference spectroscopy.

scholarly journals A Cytochrome c Oxidase Model Catalyzes Oxygen to Water Reduction Under Rate-Limiting Electron Flux

Science ◽  
2007 ◽  
Vol 315 (5818) ◽  
pp. 1565-1568 ◽  
Author(s):  
J. P. Collman ◽  
N. K. Devaraj ◽  
R. A. Decreau ◽  
Y. Yang ◽  
Y.-L. Yan ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Electron Flux ◽  
Water Reduction ◽  
Rate Limiting

scholarly journals The steady state kinetics of the NADH-dependent nitrite reductase from Escherichia coli K12. The reduction of single-electron acceptors

1982 ◽  
Vol 203 (2) ◽  
pp. 505-510 ◽  
Author(s):  
R H Jackson ◽  
J A Cole ◽  
A Cornish-Bowden
Keyword(s):  
Escherichia Coli ◽  
Cytochrome C ◽  
Nitrite Reductase ◽  
Maximum Velocity ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Steady State Kinetics ◽  
Cytochrome C Reduction ◽  
Pattern Of Variation ◽  
Rate Limiting

The kinetic characteristics of the diaphorase activities associated with the NADH-dependent nitrite reductase (EC 1.6.6.4) from Escherichia coli have been determined. The values of the apparent maximum velocity are similar for the reduction of Fe(CN)6(3)-and mammalian cytochrome c by NADH. These reactions may therefore have the same rate-limiting step. NAD+ activates NADH-dependent reduction of cytochrome c, and the apparent maximum velocity for this substrate increases more sharply with the concentration of NAD+ than for hydroxylamine. The simplest explanation is that NAD+ activation of hydroxylamine reduction derives solely from activation of steps involved in the reduction of cytochrome c, a flavin-mediated reaction, but these steps are only partly rate-limiting for the reduction of hydroxylamine. At 0.5 mM-NAD+, the apparent maximum velocity was 2.3 times higher for 0.1 mM-cytochrome c as substrate than for 100 mM-hydroxylamine, suggesting that the rate-limiting step during hydroxylamine reduction is a step that is not involved in cytochrome c reduction. A scheme is proposed that can account for the pattern of variation with [NAD+] of the Michaelis-Menten parameters for hydroxylamine and for NADH with hydroxylamine or cytochrome c as oxidized substrate.

Role of mRNA stability and translation in the expression of cytochrome c oxidase during mouse myoblast differentiation: instability of the mRNA for the liver isoform of subunit VIa

2000 ◽  
Vol 351 (1) ◽  
pp. 133-142 ◽  
Author(s):  
Elizabeth L. THAMES ◽  
Danforth A. NEWTON ◽  
Samuel A. BLACK ◽  
Lewis H. BOWMAN
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Mrna Stability ◽  
Muscle Development ◽  
Actinomycin D ◽  
Myoblast Differentiation ◽  
Element Analysis ◽  
Rate Limiting ◽  
Mouse Myoblast

The role of mRNA stability and translation in mediating the expression of selected subunits of cytochrome c oxidase (COX) was examined during the differentiation of mouse myoblasts into myotubes in cell culture. The expression of the liver (L) and heart (H) isoforms of COX VIa, which undergo an isoform switch during muscle development, as well as of the Va subunit, which is expressed in all tissues, was analysed. The translational efficiencies of COX Va, VIa-L and VIa-H, as well as of mitochondrially encoded COX mRNAs, were inferred from their distribution in polysome gradients. These experiments suggest that the translational efficiencies of these mRNAs do not change during myoblast differentiation, although the nuclear mRNAs for COX Va, VIa-L and VIa-H are translated more efficiently than the mitochondrial mRNAs. Analysis of mRNA stability using the tetracycline-repressible promoter system and/or actinomycin D indicates that COX VIa-L mRNA decays with a half-life of ∼ 5–6h in both myoblasts and myotubes, whereas COX VIa-H and Va mRNAs decay with half-lives of > 15h in myotubes. This relative instability of COX VIa-L mRNA serves to limit the accumulation of COX VIa-L mRNA in these myogenic cells, as compared with mRNAs for other COX subunits. Deletion/replacement mapping experiments suggest that the COX VIa-L 3´ untranslated region contains a destabilization element. Analysis of the rate of poly(A) tail shortening on COX VIa-L and stable α-globin mRNAs suggests that the overall rate of poly(A) shortening per se is not rate limiting for the degradation of COX VIa-L mRNA.

scholarly journals Block of current through single calcium channels by Fe, Co, and Ni. Location of the transition metal binding site in the pore.

1991 ◽  
Vol 97 (2) ◽  
pp. 351-367 ◽  
Author(s):  
B D Winegar ◽  
R Kelly ◽  
J B Lansman
Keyword(s):  
Metal Binding ◽  
The Other ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Coulombic Interactions ◽  
Rate Limiting Step ◽  
Ion Size ◽  
Rate Limiting ◽  
Kinetics Of ◽  
Ca2 Channels

The blocking actions of Fe2+, Co2+, and Ni2+ on unitary currents carried by Ba2+ through single dihydropyridine-sensitive Ca2+ channels were recorded from cell-attached patches on myotubes from the mouse C2 cell line. Adding millimolar concentrations of blocker to patch electrodes containing 110 mM BaCl2 produced discrete excursions to the closed channel level. The kinetics of blocking and unblocking were well described with a simple model of open channel block. Hyperpolarization speeded the exit of all of the blockers from the channel, as expected if the blocking site resides within the pore. The block by Ni2+ differs from that produced by Fe2+ and Co2+ because Ni2+ enters the channel approximately 20 times more slowly and exits approximately 50 times more slowly. Ni2+ also differs from the other transition metals because at millimolar concentrations it reduces the amplitude of the unitary current in a concentration-dependent manner. The results are consistent with the idea that the rate-limiting step for ion entry into the channel is water loss at its inner coordination sphere; unblocking, on the other hand, cannot be explained in terms of simple coulombic interactions arising from differences in ion size.

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