A comparison of nitric oxide and hydrogen sulphide interactions with mitochondrial cytochrome c oxidase

Mitochondrial cytochrome c oxidase (CCO) is responsible for >95% of mammalian oxygen consumption, and is key to generation of the electrochemical proton gradient required for the bulk of ATP production. Nitric oxide (NO) is a signalling molecule with roles including the control of blood flow and blood pressure via activation of soluble guanylate cyclase. Hydrogen sulphide (H2S) has been proposed to be a signalling molecule regulating, amongst other things, vascular tone and myocardial contractility through activation of ATP-sensitive potassium channels. NO and H2S both have complex interactions with CCO, resulting in inhibition of enzyme activity and/or metabolism of the gas itself. It has therefore been proposed that CCO may play an important role in NO and/or H2S signal transduction.In order to determine the importance of these interactions in vivo it is vital to evaluate the concentrations where they exert their effects. We used polarographic and spectroscopic tools to determine simultaneously the metabolism of the gas, enzyme activity and the functional state of the enzyme. We combined these data with kinetic modelling to report on reaction routes and rates. Data were consistent with an interaction of H2S with oxidised CCO, with on and off rate constants of 3700 M?1 s?1 and 1.4 � 10?1 s?1, respectively. The resulting dissociation constant, Kd = 0.04 ?M, suggests an oxygen-insensitive IC50(H2S) of 53 ?M; comparisons with NO under identical conditions suggest that, molecule for molecule, H2S is over 1000 less effective a CCO inhibitor than NO under physiological conditions. The implications for this finding are discussed.