Functional Dissection of the Proton Pumping Modules of Mitochondrial Complex I

Abstract

Author SummaryMitochondria—the power plants of eukaryotic cells—produce energy in the form of ATP. More than one-third of this energy production is driven by a gradient of protons across the mitochondrial membrane created by the pumping action of a very large enzyme called complex I. Defects in complex I are implicated in numerous pathological processes like neurodegeneration and biological aging. Recent X-ray structural analyses revealed that complex I is an L-shaped molecule with one arm integrated into the membrane and the other sticking into the aqueous interior of the mitochondrion; the chemical reactions of the enzyme take place in this hydrophilic arm, clearly separated from proton pumping that must occur somewhere in the membrane arm. To assign the pump function to structural domains, we created a stable subcomplex of complex I by deleting the gene encoding one of its small subunits in a yeast called Yarrowia lipolytica. This subcomplex lacked half of the membrane arm; it was still catalytically active but it pumped only half the number of protons as the full complex. This indicates that complex I has two functionally distinct pump modules operating in its membrane arm.