Abstract
Thyroid hormones regulate tissue metabolism to establish an energy balance in the cell, in particular, by affecting oxidative phosphorylation. Their long-term impact is mainly associated with changes in gene expression, while the short-term effects may differ in their mechanisms. Our work was devoted to studying the short-term effects of hormones T2, T3 and T4 on mitochondrial cytochrome c oxidase (CcO) mediated by direct contact with the enzyme. The data obtained indicate the existence of two separate sites of CcO interaction with thyroid hormones, differing in their location, affinity and specificity to hormone binding. First, we show that T3 and T4 but not T2 inhibit the oxidase activity of CcO in solution and on membrane preparations with Ki ≈ 100–200 μM. In solution, T3 and T4 compete in a 1:1 ratio with the detergent dodecyl-maltoside to bind to the enzyme. The peroxidase and catalase partial activities of CcO are not sensitive to hormones, but electron transfer from heme a to the oxidized binuclear center is affected. We believe that T3 and T4 could be ligands of the bile acid-binding site found in the 3D structure of CcO by Ferguson-Miller’s group, and hormone-induced inhibition is associated with dysfunction of the K-proton channel. A possible role of this interaction in the physiological regulation of the enzyme is discussed. Second, we find that T2, T3, and T4 inhibit superoxide generation by oxidized CcO in the presence of excess H2O2. Inhibition is characterized by Ki values of 0.3–5 μM and apparently affects the formation of O2●− at the protein surface. The second binding site for thyroid hormones presumably coincides with the point of tight T2 binding on the Va subunit described in the literature.
Highlights
Cytochrome c oxidase (CcO) is the terminal enzyme of the respiratory chain located in the coupling membranes of mitochondria and aerobic bacteria
As we have found recently, a number of steroid-like compounds have a pronounced inhibitory effect on solubilized CcO from mitochondria [14,15]
We assumed that steroid-like inhibitors may be ligands of the bile acid-binding site (BABS) site and their inhibitory effect associated with impaired conductivity of the proton channel K
Summary
Cytochrome c oxidase (CcO) (for recent reviews, see [1,2]) is the terminal enzyme of the respiratory chain located in the coupling membranes of mitochondria and aerobic bacteria. The catalytic center of the enzyme is a binuclear center a3/CuB formed by closely spaced high-spin heme a3 and copper ion. Electrons arriving from cytochrome c are transferred along the redox centers: CuA → low-spin heme a → a3/CuB → O2. The four-electron reduction of oxygen to water in the binuclear center is coupled to proton translocation and energy transduction to ∆μH+. Mitochondrial CcO consists of 13 subunits [3], the three largest of which (catalytic subunit I containing heme a, a3/CuB binuclear center and the proton channels, subunit II carrying cytochrome c binding site and CuA and subunit III) are homologous to the subunits of bacterial enzymes and encoded by mitochondrial DNA. The K-channel is closed and protons are transferred across the membrane through channel D (for details, see reviews [6–8])
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