Cytochrome c oxidase inhibition by calcium at physiological ionic composition of the medium: Implications for physiological significance of the effect.

Cytochrome c oxidase (CcO) from mammalian mitochondria binds Ca and Na in a special cation binding site. Binding of Ca brings about partial inhibition of the enzyme while Na competes with Ca for the binding site and protects the enzyme from the inhibition [Vygodina, T., Kirichenko, A. and Konstantinov, A.A. (2013). Direct Regulation of Cytochrome c oxidase by Calcium Ions. PLoS One 8(9): e74436]. In the original studies, the inhibition was found to depend significantly on the ionic composition of the buffer. Here we describe inhibition of CcO by Ca in media containing the main ionic components of cytoplasm (150mM KCl, 12mM NaCl and 1mM MgCl). Under these conditions, Ca inhibits CcO with effective K of 20-26μM, that is an order of magnitude higher than determined earlier in the absence of Na. At physiological value of ionic strength, the inhibition can be observed at any turnover number of CcO, rather than only at low TN (<10s) as found previously. The inhibition requires partially oxidized state of cytochrome c and is favored by high ionic strength with a sharp transition at 0.1-0.2M. The high K=20-26μM found for CcO inhibition by calcium matches closely the known value of "K" for Ca-induced activation of the mitochondrial calcium uniporter. The inhibition of CcO by Ca is proposed to modulate mitochondrial Ca-uptake via the mitochondrial calcium uniporter, promote permeability transition pore opening and induce reduction of Mia40 in the mitochondrial intermembrane space.