Abstract

Cells depend on a continuous supply of ATP, the universal energy currency. In mitochondria, ATP is produced by a series of redox reactions, whereby an electrochemical gradient is established across the inner mitochondrial membrane. The ATP synthase harnesses the energy of the gradient to generate ATP from ADP and inorganic phosphate. We determined the structure of ATP synthase within mitochondria of the unicellular alga Polytomella by electron cryo-tomography. Sub-tomogram averaging revealed six rotary positions of the central stalk, subclassified into 21 substates of the F1 head. The Polytomella ATP synthase forms helical arrays with multiple adjacent rows defining the cristae ridges. The structure of ATP synthase under native operating conditions in the presence of a membrane potential represents a pivotal step toward the analysis of membrane protein complexes in situ.