Investigation of the Lipid Dependence of Respiratory Complex IV Activation Using Nanoscale Bilayers

The mitochondrion’s energy-transducing inner membrane is home to the electron transport chain, a meshwork of protein complexes vital to maintaining cellular energy balance. The unique lipid composition of the inner membrane of the mitochondrion includes the dimeric phospholipid, cardiolipin, which is vital for the efficient activity of respiratory Complex IV, the terminal member of the electron transport chain. Previous studies have investigated the protein-lipid interaction between respiratory Complex IV and cardiolipin in detergent micelles and found endogenous cardiolipin molecules, which co-purified with the protein. In this study, respiratory Complex IV was purified and the endogenous cardiolipin molecules were removed with porcine phospholipase A2, before being incorporated into the biologically relevant nanodisc model membrane system. The cardiolipin-free respiratory Complex IV was reconstituted into a controlled lipid environment containing the bilayer favoring phospholipid POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) or a biomimetic mixture of lipids, including POPC, POPE (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine) and tetraoleoyl cardiolipin, in molar ratios similar to the lipid environment found in the inner mitochondrial membrane. Using this model system, it has been shown that cardiolipin is vital for the proper activity of respiratory Complex IV. Moreover, this work provides initial evidence that cardiolipin contributes to the activity of Complex IV through the presence bulk lipids that do not directly interface with Complex IV as annular lipids. These findings are significant, as they provide insight into the pathology of diseases associated with mitochondrial lipid remodeling pathways such as Barth Syndrome