Structural studies on bovine heart cytochrome c oxidase

Area of Science:

• Biochemistry
• Structural Biology
• Molecular Biophysics

Background:

• Cytochrome c oxidase (CcO) is a crucial enzyme in cellular respiration.
• Previous X-ray structures of bovine heart CcO provided insights into its complex composition, including protein subunits, lipids, and metal sites.

Purpose of the Study:

• To elucidate the structural mechanisms of oxygen reduction and proton transport in bovine heart CcO.
• To investigate the role of specific sites and pathways in enzyme function.

Main Methods:

• High-resolution X-ray crystallography (up to 1.8Å resolution).
• Analysis of oxygen analog binding.
• Site-directed mutagenesis of bovine CcO expressed in HeLa cells.

Main Results:

• Oxygen molecules transiently bind at the Cu(B) site before reduction by Fe(a3), forming O(2)(-).
• Three electron transfer pathways (from Cu(B), Fe(a3), and Tyr244) facilitate the non-sequential 3-electron reduction of O(2)(-), breaking the OO bond without releasing active oxygen species.
• A proton conducting pathway, comprising a hydrogen-bond network and a water channel, facilitates proton transport between cellular compartments.
• Heme a oxidation delocalizes positive charge to heme peripheral groups, driving proton transport.
• Mutagenesis confirmed the roles of the proton pathway and its blocking of reverse proton transfer.

Conclusions:

• Bovine heart CcO employs sophisticated electron and proton transfer mechanisms for efficient energy conversion.
• The identified pathways and structural features are critical for preventing the release of reactive oxygen species during respiration.
• Mutagenesis studies validate the functional significance of the proton channel and its regulatory elements.