Mitochondrial ATP synthasome. Cristae-enriched membranes and a multiwell detergent screening assay yield dispersed single complexes containing the ATP synthase and carriers for Pi and ADP/ATP.
The terminal step of ATP synthesis in intact mitochondria is catalyzed by the ATP synthase (F(0)F(1)) that works in close synchrony with the P(i) and ADP/ATP carriers. Each carrier consists of only a single polypeptide chain in dimeric form, while the ATP synthase is highly complex consisting in animals of 17 known subunit types and more than 30 total subunits. Although structures at high resolution have been obtained for the water-soluble F(1) part of the ATP synthase consisting of only five subunit types, such structures have not been obtained for either the complete ATP synthase or the P(i) and ADP/ATP carriers. Here, we report that all three proteins are localized in highly purified cristae-like vesicles obtained by extensive subfractionation of the mitochondrial inner membrane. Moreover, using a multiwell detergent screening assay, 4 nonionic detergents out of 80 tested were found to disperse these cristae-like vesicles into single soluble complexes or "ATP synthasomes" that contain the ATP synthase in association with the P(i) and ADP/ATP carriers. These studies offer new mechanistic insights into the terminal steps of oxidative phosphorylation in mitochondria and set the stage for future structural efforts designed to visualize in atomic detail the entire complex involved. They also provide evidence that the cristae are a subcompartment of the inner membrane.
Ko
Young H
YH
Department of Biological Chemistry, The Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA.
Delannoy
Michael
M
Hullihen
Joanne
J
Chiu
Wah
W
Pedersen
Peter L
PL
D000818:Animals; D015153:Blotting, Western; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D007425:Intracellular Membranes / Q000201:enzymology; D008297:Male; D008854:Microscopy, Electron; D008928:Mitochondria / Q000201:enzymology*; D025261:Mitochondrial Proton-Translocating ATPases / Q000302:isolation & purification* / Q000378:metabolism / Q000648:ultrastructure; D009840:Oligomycins; D051381:Rats; D017207:Rats, Sprague-Dawley; D020539:Sequence Analysis, Protein; D013347:Subcellular Fractions / Q000201:enzymology; D014475:Uncoupling Agents
D003902:Detergents; D009840:Oligomycins; D014475:Uncoupling Agents; D025261:Mitochondrial Proton-Translocating ATPases