Effect of Calcium Ions on Electron Transfer between Hemes a and a₃ in Cytochrome c Oxidase

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T. V. Vygodina^*, A. V. Dyuba, and A. A. Konstantinov

Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119991 Moscow, Russia; E-mail: vygodina@genebee.msu.ru

^* To whom correspondence should be addressed.

Received February 28, 2012; Revision received March 15, 2012

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Kinetics of the reduction of the hemes in cytochrome c oxidase in the presence of high concentration of ruthenium(III)hexaammine chloride was examined using a stopped-flow spectrophotometer. Upon mixing of the oxidized enzyme with dithionite and Ru(NH₃)₆³⁺, three well-resolved phases were observed: heme a reduction reaching completion within a few milliseconds is followed by two slow phases of heme a₃ reduction. The difference spectrum of heme a₃ reduction in the visible region is characterized by a maximum at ~612 nm, rather than at 603 nm as was believed earlier. It is shown that in the case of bovine heart cytochrome c oxidase containing a special cation-binding site in which reversible binding of calcium ion occurs, heme a₃ reduction is slowed down by low concentrations of Ca²⁺. The effect is absent in the case of the bacterial cytochrome oxidase in which the cation-binding site contains a tightly bound Ca²⁺ ion. The data corroborate the inhibition of the cytochrome oxidase enzymatic activity by Ca²⁺ ions discovered earlier and indicate that the cation affects intramolecular electron transfer.

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KEY WORDS: cytochrome c oxidase, Ca²⁺ ions, fast kinetics, heme a₃

DOI: 10.1134/S0006297912080111

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