title: Resolving stepping rotation in Thermus thermophilus H(+)-ATPase/synthase with an essentially drag-free probe.
abstract: Vacuole-type ATPases (V(o)V₁) and F(o)F₁ ATP synthases couple ATP hydrolysis/synthesis in the soluble V(1) or F₁ portion with proton (or Na(+)) flow in the membrane-embedded V(o) or F(o) portion through rotation of one common shaft. Here we show at submillisecond resolutions the ATP-driven rotation of isolated V₁ and the whole V(o)V₁ from Thermus thermophilus, by attaching a 40-nm gold bead for which viscous drag is almost negligible. V₁ made 120° steps, commensurate with the presence of three catalytic sites. Dwells between the steps involved at least two events other than ATP binding, one likely to be ATP hydrolysis. V(o)V₁ exhibited 12 dwell positions per revolution, consistent with the 12-fold symmetry of the V(o) rotor in T. thermophilus. Unlike F₁ that undergoes 80°-40° substepping, chemo-mechanical checkpoints in isolated V₁ are all at the ATP-waiting position, and V(o) adds further bumps through stator-rotor interactions outside and remote from V₁.
authors.affiliation: Department of Physics, Faculty of Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan.
authors.forename: Shou
authors.initials: S
authors.lastname: Furuike
authors.forename: Masahiro
authors.initials: M
authors.lastname: Nakano
authors.forename: Kengo
authors.initials: K
authors.lastname: Adachi
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Noji
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Kinosita
authors.forename: Ken
authors.initials: K
authors.lastname: Yokoyama
mesh_terms: D000255:Adenosine Triphosphate / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020134:Catalytic Domain; D004926:Escherichia coli; D006046:Gold / Q000032:analysis / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D008863:Microspheres; D008958:Models, Molecular; D015335:Molecular Probes / Q000032:analysis / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011522:Protons; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012399:Rotation; D012964:Sodium / Q000378:metabolism; D019809:Streptavidin / Q000032:analysis / Q000378:metabolism; D016963:Thermus thermophilus / Q000737:chemistry / Q000201:enzymology / Q000235:genetics; D025262:Vacuolar Proton-Translocating ATPases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014783:Viscosity
chemical_list: D001426:Bacterial Proteins; D015335:Molecular Probes; D021122:Protein Subunits; D011522:Protons; D011994:Recombinant Proteins; D006046:Gold; D000255:Adenosine Triphosphate; D019809:Streptavidin; D012964:Sodium; D025262:Vacuolar Proton-Translocating ATPases