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Movements of the ε‐subunit during catalysis and activation in single membrane‐bound H + ‐ATP synthase
Author(s) -
Zimmermann Boris,
Diez Manuel,
Zarrabi Nawid,
Gräber Peter,
Börsch Michael
Publication year - 2005
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1038/sj.emboj.7600682
Subject(s) - atp synthase , förster resonance energy transfer , chemistry , protein subunit , chemiosmosis , conformational change , enzyme , atp hydrolysis , electron transport chain , stereochemistry , active site , biophysics , f atpase , atp synthase gamma subunit , chloroplast , biochemistry , fluorescence , atpase , biology , thylakoid , physics , quantum mechanics , gene
F 0 F 1 ‐ATP synthases catalyze proton transport‐coupled ATP synthesis in bacteria, chloroplasts, and mitochondria. In these complexes, the ε‐subunit is involved in the catalytic reaction and the activation of the enzyme. Fluorescence‐labeled F 0 F 1 from Escherichia coli was incorporated into liposomes. Single‐molecule fluorescence resonance energy transfer (FRET) revealed that the ε‐subunit rotates stepwise showing three distinct distances to the b ‐subunits in the peripheral stalk. Rotation occurred in opposite directions during ATP synthesis and hydrolysis. Analysis of the dwell times of each FRET state revealed different reactivities of the three catalytic sites that depended on the relative orientation of ε during rotation. Proton transport through the enzyme in the absence of nucleotides led to conformational changes of ε. When the enzyme was inactive (i.e. in the absence of substrates or without membrane energization), three distances were found again, which differed from those of the active enzyme. The three states of the inactive enzyme were unequally populated. We conclude that the active–inactive transition was associated with a conformational change of ε within the central stalk.