Modeling the detailed kinetics of mitochondrial cytochromecoxidase: Catalytic mechanism and nitric oxide inhibition

Abstract

Cytochrome c oxidase (CcO) catalyzes the exothermic reduction of O to HO by using electrons from cytochrome c, and hence plays a crucial role in ATP production. Although details on the enzyme structure and redox centers involved in O reduction have been known, there still remains a considerable ambiguity on its mechanism of action, e.g., the number of sequential electrons donated to O in each catalytic step, the sites of protonation and proton pumping, and nitric oxide (NO) inhibition mechanism. In this work, we developed a thermodynamically constrained mechanistic mathematical model for the catalytic action of CcO based on available kinetic data. The model considers a minimal number of redox centers on CcO and couples electron transfer and proton pumping driven by proton motive force (PMF), and accounts for the inhibitory effects of NO on the reaction kinetics. The model is able to fit well all the available kinetic data under diverse experimental conditions with a physiologically realistic unique parameter set. The model predictions show that: 1) the apparent K of O varies considerably and increases from fully reduced to fully oxidized cytochrome c depending on pH and the energy state of mitochondria, and 2) the intermediate enzyme states depend on pH and cytochrome c redox fraction and play a central role in coupling mitochondrial respiration to PMF. The developed CcO model can easily be integrated into existing mitochondrial bioenergetics models to understand the role of the enzyme in controlling oxidative phosphorylation in normal and disease conditions.

Isolation of mitochondrial subpopulations from skeletal muscle: Optimizing recovery and preserving integrity

“…Also, in a rat skeletal muscle study, a twofold increase in the respiratory capacity in white, red, and mixed muscle fibres was attributed to an approximately twofold increase of citrate synthase, cytochrome c , and cytochrome c oxidase . Computational studies confirmed a close relationship among cytochrome c reduced and oxidized concentrations and mitochondrial respiration rate …”

“…The reduced content of cytochrome c and c 1 found in SSM in comparison to that observed for IFM is consistent with a lower respiration rate of SSM compared to that of IFM. Experimental and computational studies support a strong relationship between cytochrome c content and respiratory capacity. In a rat kidney study from our laboratory, a decrease in oxidative phosphorylation was reported to be related to a reduced cytochrome c content of mitochondria with intact outer membranes .…”

The Effect of Mitochondrial Complex I-Linked Respiration by Isoflurane Is Independent of Mitochondrial Nitric Oxide Production

“…NO acts as a multisite inhibitor of the mitochondrial ETC. The most sensitive and widely studied target for NO in mitochondria is the terminal enzyme of the ETC, cytochrome c oxidase [9, 23, 24]. NO competes with O 2 at the binuclear Cu B /cytochrome a 3 site.…”

“…NO competes with O 2 at the binuclear Cu B /cytochrome a 3 site. This competition with O 2 results in the inhibition of the enzyme, suggesting that NO is an important physiological regulator of mitochondrial oxidative phosphorylation [9, 23, 24]. Reports [25, 26] that the presence of the mitochondrial NOS isoenzyme is a constitutive protein in the mitochondrial inner membrane support a potential physiological role for NO in mitochondrial respiration.…”

NO control of mitochondrial function in normal and transformed cells

“…The most important sensitive site to NO is COX, which is the terminal electron acceptor of the respiratory chain and is the enzyme complex that reduces molecular oxygen to H 2 O, helping generate a proton gradient that drives Complex V and ATP synthesis. The kind of NO inhibition is dependent on COX redox state (52–54). COX is reduced after receiving electrons from cytochrome c and the reduction of the metals (Fe2+ and Cu+) when molecular oxygen binds to a heme-copper binuclear active site (53).…”