scholarly journals Kinetic studies on the binding of cyanide to oxygenated cytochrome c oxidase

1976 ◽  
Vol 155 (2) ◽  
pp. 453-455 ◽  
Author(s):  
T Brittain ◽  
C Greenwood
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Kinetic Studies ◽  
Binding Properties ◽  
Cyanide Concentration ◽  
Cyanide Binding ◽  
Single Process ◽  
Reduced Forms

The reaction of cyanide with oxygenated cytochrome c oxidase was followed by means of flow-flash techniques. The oxygenated form, produced after photolysis of the partially reduced CO complex in the presence of cyanide and O2, shows cyanide-binding properties distinct from those of both the oxidized and the reduced forms of the protein. The binding is a single process (k = 22M-1-S-1) linearly dependent on cyanide concentration to as high as 75 mM. It is suggested that the oxygenated form is a conformational variant of the oxidized protein.

scholarly journals A re-examination of the reactions of cyanide with cytochrome c oxidase

1984 ◽  
Vol 220 (1) ◽  
pp. 57-66 ◽  
Author(s):  
M G Jones ◽  
D Bickar ◽  
M T Wilson ◽  
M Brunori ◽  
A Colosimo ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Essential Feature ◽  
Tight Binding ◽  
Sodium Dithionite ◽  
Difference Spectra ◽  
Ferrocytochrome C ◽  
Cyanide Binding ◽  
Kinetic Predictions ◽  
Reduced Forms

Experiments were performed to examine the cyanide-binding properties of resting and pulsed cytochrome c oxidase in both their stable and transient turnover states. Inhibition of the oxidation of ferrocytochrome c was monitored as a function of cyanide concentration. Cyanide binding to partially reduced forms produced by mixing cytochrome c oxidase with sodium dithionite was also examined. A model is presented that accounts fully for cyanide inhibition of the enzyme, the essential feature of which is the rapid, tight, binding of cyanide to transient, partially reduced, forms of the enzyme populated during turnover. Computer fitting of the experimentally obtained data to the kinetic predictions given by this model indicate that the cyanide-sensitive form of the enzyme binds the ligand with combination constants in excess of 10(6) M-1 X s-1 and with KD values of 50 nM or less. Kinetic difference spectra indicate that cyanide binds to oxidized cytochrome a33+ and that this occurs rapidly only when cytochrome a and CuA are reduced.

Kinetic Studies on Mammalian Cytochrome c Oxidase

1973 ◽  
Vol 1 (1) ◽  
pp. 34-35 ◽  
Author(s):  
E. ANTONINI ◽  
M. BRUNORI ◽  
C. GREENWOOD ◽  
M. T. WILSON
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Kinetic Studies

scholarly journals Transient kinetic studies of Neurospora crassa cytochrome c oxidase

1983 ◽  
Vol 215 (2) ◽  
pp. 425-427 ◽  
Author(s):  
M Brunori ◽  
M C Silvestrini ◽  
M T Wilson ◽  
H Weiss
Keyword(s):  
Cytochrome C ◽  
Neurospora Crassa ◽  
Cytochrome C Oxidase ◽  
Flash Photolysis ◽  
Kinetic Studies ◽  
Dissociation Rate ◽  
Steady State Kinetic ◽  
Kinetic Investigations ◽  
Dissociation Rate Constants

The reaction of Neurospora crassa cytochrome c oxidase with CO was studied by flash-photolysis and rapid-mixing experiments, leading to the determination of the association and dissociation rate constants (7 X 10(4) M-1 X s-1 and 0.02s-1 respectively). Pre-steady-state kinetic investigations of the catalytic properties of the enzyme showed that under proper conditions Neurospora cytochrome c oxidase can be ‘pulsed’, i.e. activated, like the mammalian enzyme. The ‘pulsed’ species is spectroscopically different from the ‘resting’ one, and the decay into the ‘resting’ state is fast (t1/2 approx. 3 min).

scholarly journals Kinetic studies on the reaction between cytochrome c oxidase and ferrocytochrome c

1975 ◽  
Vol 147 (1) ◽  
pp. 145-153 ◽  
Author(s):  
M T Wilson ◽  
C Greenwood ◽  
M Brunori ◽  
E Antonini
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Time Course ◽  
Kinetic Studies ◽  
Initial Reaction ◽  
Fast Phase ◽  
Ferrocytochrome C ◽  
Cytochrome C2 ◽  
Kinetics Of ◽  
Flow Experiments

In stopped-flow experiments in which oxidized cytochrome c oxidase was mixed with ferrocytochrome c in the presence of a range of oxygen concentrations and in the absence and presence of cyanide, a fast phase, reflecting a rapid approach to an equilibrium, was observed. Within this phase, one or two molecules of ferrocytochrome were oxidized per haem group of cytochrome a, depending on the concentration of ferrocytochrome c used. The reasons for this are discussed in terms of a mechanism in which all electrons enter through cytochrome a, which, in turn, is in rapid equilibrium with a second site, identified with ‘visible’ copper (830 nm-absorbing) Cud (Beinert et al., 1971). The value of the bimolecular rate constant for the reaction between cytochromes c2+ and a3+ was between 10(6) and 10(7) M(-1)-S(-1); some variability from preparation to preparation was observed. At high ferrocytochrome c concentrations, the initial reaction of cytochrome c2+ with cytochrome a3+ could be isolated from the reaction involving the ‘visible’ copper and the stoicheiometry was found to approach one molecule of cytochrome c2+ oxidized for each molecule of cytochrome a3+ reduced. At low ferrocytochrome c concentrations, however, both sites (i.e. cytochrome a and Cud) were reduced simultaneously and the stoicheiometry of the initial reaction was closer to two molecules of cytochrome c2+ oxidized per molecule of cytochrome a reduced. The bleaching of the 830 nm band lagged behind or was simultaneous with the formation of the 605 nm band and does not depend on the cytochrome c concentration, whereas the extinction at the steady-state does. The time-course of the return of the 830 nm-absorbing species is much faster than the bleaching of the 605 nm-absorbing component, and parallels that of the turnover phase of cytochrome c2+ oxidation. Additions of cyanide to the oxidase preparations had no effect on the observed stoicheiometry or kinetics of the reduction of cytochrome a and ‘visible’ copper, but inhibited electron transfer to the other two sites, cytochrome a3 and the undetectable copper, Cuu.

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