Characterization of the Electron Transport Chain Metabolon Bioelectrocatalysis

The electron transport chain is a group of membrane redox enzymes that are the driving force for the production of energy in mitochondria. Several theories and models have circulated different structures of the enzymes in the inner membrane.¹ The three enzyme complexes form a supercomplex or respirasome, where Complex I, dimer Complex III, and various copies of Complex IV are electrostatically, hydrophobically and catalytically connected. While structural, kinetic, and genetic evidence prove the formation of the supercomplex, characterization of the bioelectrocatalysis has never been attempted of the three enzymes together on an electrode.² The supercomplex contains multiple redox cofactors of the individual complex that span the lipid membrane and can come in contact with the electrode surface. By purifying the electron transport chain supercomplex, we have been able to reconstruct the inner membrane and immobilizing it onto a gold electrode. A polymer tethered lipid bilayer allows for the study of the natural bioelectrocatalysis of the large transmembrane proteins. With the addition of oxidized cytochrome c, Complex IV gives a catalytic response indicating that Complex III and Complex IV are active. With the addition of NADH and oxidized cytochrome c, the catalytic peaks increase. This indicates that all three enzymes are working together increasing their redox activities at the surface of the electrode. The supercomplex is further confirmed by inhibition of CIII and CIV, eliminating all activity of the enzymes at the electrode surface. This evidence shows how dependent the enzymes are on each other for efficient catalytic activity. The characterization of the supercomplex of these transmembrane redox enzymes and their bioelectrocatalysis on the electrode is the first for studying a natural metabolon electrochemically.

 References

1.         Hackenbrock, C. R.; Chazotte, B.; Gupte, S. S., The random collision model and a critical assessment of diffusion and collision in mitochondrial electron transport. J Bioenerg Biomembr 1986, 18 (5), 331-368.

2.         (a) Genova, M. L.; Lenaz, G., Functional role of mitochondrial respiratory supercomplexes. Biochimica et Biophysica Acta (BBA) - Bioenergetics 2014, 1837 (4), 427-443; (b) Genova, M. L.; Baracca, A.; Biondi, A.; Casalena, G.; Faccioli, M.; Falasca, A. I.; Formiggini, G.; Sgarbi, G.; Solaini, G.; Lenaz, G., Is supercomplex organization of the respiratory chain required for optimal electron transfer activity? Biochimica et Biophysica Acta (BBA) - Bioenergetics 2008, 1777 (7-8), 740-746.