Cytochrome c oxidase
Mammalian CcO has a molecular weight of about 200 kDa and contains 13 different polypeptide subunits. The three largest form the core of the enzyme and are encoded by the mitochondrial genome, whereas the remaining 10 subunits originate from nuclear DNA. Bacterial CcO are simpler in structure and have only from three to four subunits, but the sequence homology of subunits I, II, and III to the corresponding ones of mitochondrial CcO is extremely high.
At present, mitochondrial and bacterial oxidases from five organisms have been resolved with resolution up to 1.8 Å.
Subunit I is the largest subunit of CcO with a molecular weight of about 60 kDa. It consists of 12 transmembrane helices without any large extramembrane domain When viewed from the top (P-side), the 12 segments of subunit I form three semicircular arcs with three pores in the centers of the arcs.
The second A-type heme is situated at about 13 Å (center-to-center distance) from heme a and is denoted as a3. Heme a3 is a high-spin heme which means that the heme has only one histidine ligand leaving one side of the heme empty and available for binding of ligands such as dioxygen, carbon monoxide, water molecules etc.
The last redox metal center of subunit I is ~5 Å away from heme a3 iron and is formed by a copper atom denoted as CuB. Together, heme a3 and CuB form the binuclear catalytic center of the oxidase responsible for binding and subsequent catalysis of oxygen.
In addition to the redox-active centers, subunit I also contains tightly bound non-redox active metal centers: a Mg2+/Mn2+ binding site at the interface between subunits I and II, and a sodium (in mitochondria) or calcium (in bacteria) binding site in a loop between helices I and II close to the P-side of the membrane.
Subunit II is another CcO subunit, which contains redox active cofactors. It has a molecular weight of about 27 kDa, and forms two transmembrane helices interacting with subunit I and a large C-terminal hydrophilic globular domain at the P-side of the membrane. The redox active copper center situated in the globular domain almost on the border with subunit I, conventionally referred to as CuA, is formed by two copper atoms. The CuA center in the oxidized state of the enzyme was found to be in mixed-valence configuration that can be formally represented as [Cu1.5+-Cu1.5+]. Upon reduction CuA holds an electron by sharing it between both copper atoms.
Subunit III is the biggest subunit of CcO that has no redox cofactors. It has a molecular weight of about 30 kDa, and consists of seven transmembrane helices without any extensive extramembrane domain. The helices of subunit III are arranged into two bundles. The bundles of helices are tilted with respect to each other forming a large V-shaped cleft joined at the N-side of the membrane. Subunit III might be involved in the stabilization of the mature oxidase and in ensuring correct assembly of the enzyme. In the same time, the V-shaped cleft is located at the mouth of the oxygen conducting channel and may secure a constant flux of oxygen into the catalytic center. It is also possible that the membrane-anchored cytochrome c552, which is a physiological electron donor for CcO from P. denitrificans , might use this cleft for binding and placing itself in an appropriate position for electron transfer to CuA.
In addition to the three core subunits, heme-copper terminal oxidases can have extra subunits. CcO from P. denitrificans has one additional subunit with a molecular weight of about 5 kDa. It has one transmembrane helix, which is in contact with all other subunits.
In mammalian CcO the three mitochondrially-encoded core subunits are supplemented by ten additional subunits, which are encoded by nuclear DNA. Seven out of ten nuclear encoded subunits consist of one transmembrane helix each, while subunits Va, Vb at the N-side and VIb at the P-side are small globular proteins. Subunit Vb contains tightly bound Zn2+.
© Ilya Belevich & Michael Verkhovsky