Methods to determine the status of mitochondrial ATP synthase assembly.
The adenosine triphosphate (ATP) synthase (F1-F0 complex) of the mitochondrial inner membrane is responsible for making nearly all of the ATP utilized by eukaryotic organisms. The enzyme is an oligomer of more than 20 different subunits, 14 of which are essential for its catalytic activity. The other subunits function in the regulation and structure of the complex. Subunits essential for catalytic activity make up the proton pore, the bulk of the F1 headpiece, and the two stalks that physically and functionally couple the catalytic and proton-translocating activities of the ATP synthase. Saccharomyces cerevisiae provides an excellent model system for studying mutations that affect assembly of the complex because of the ability of this organism to survive on the ATP produced from fermentation in the absence of mitochondrial respiration or oxidative phosphorylation. Studies of such mutants have been instrumental in identifying novel molecular chaperones that act at discrete steps of F1-F0 assembly. Here, we describe some experimental approaches useful in assessing the status of F1-F0 assembly.
Ackerman
Sharon H
SH
Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, Detroit, USA.
Tzagoloff
Alexander
A
D000255:Adenosine Triphosphate / Q000378:metabolism; D015153:Blotting, Western; D002499:Centrifugation, Density Gradient; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D008928:Mitochondria / Q000201:enzymology; D025261:Mitochondrial Proton-Translocating ATPases / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D008967:Molecular Biology / Q000379:methods*; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D010710:Phosphates / Q000378:metabolism; D021122:Protein Subunits / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D012995:Solubility; D013010:Sonication; D013053:Spectrophotometry; D013367:Submitochondrial Particles / Q000378:metabolism
D010710:Phosphates; D021122:Protein Subunits; D009243:NAD; D000255:Adenosine Triphosphate; D025261:Mitochondrial Proton-Translocating ATPases