Nature Communications | Article Open

Resolving stepping rotation in Thermus thermophilus H+-ATPase/synthase with an essentially drag-free probe

Shou Furuike1, 2, Masahiro Nakano3, 4, Kengo Adachi1, 7, Hiroyuki Noji4, Kazuhiko Kinosita Jr1, Ken Yokoyama3, 5, 6
Journal name:  Nature Communications
Volume:  2
Article number:  233
DOI:doi:  10.1038/ncomms1215
Received 
Accepted 
Published

Abstract

Vacuole-type ATPases (VoV1) and FoF1 ATP synthases couple ATP hydrolysis/synthesis in the soluble V1 or F1 portion with proton (or Na+) flow in the membrane-embedded Vo or Fo portion through rotation of one common shaft. Here we show at submillisecond resolutions the ATP-driven rotation of isolated V1 and the whole VoV1 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. VoV1 exhibited 12 dwell positions per revolution, consistent with the 12-fold symmetry of the Vo 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 Vo adds further bumps through stator–rotor interactions outside and remote from V1.

Affiliations

  1. Department of Physics, Faculty of Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 169-8555, Japan.
  2. Department of Physics, Osaka Medical College, Takatsuki, Osaka 569-8686, Japan.
  3. Chemical Resources Laboratory, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8503, Japan.
  4. Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka 567-0047, Japan.
  5. ICORP, ATP Synthesis Regulation Project, Japan Science and Technology Agency (JST), National Museum of Emerging Science and Innovation, 2-41 Aomi, Koto-ku, Tokyo 135-0064, Japan.
  6. Department of Molecular Biosciences, Kyoto Sangyo University, Motoyama Kamigamo, Kita-ku, Kyoto 603-8555, Japan.
  7. Present address: Department of Physics, Faculty of Science, Gakushuin University, Toshima-ku, Tokyo 171-8588, Japan.