Stepping rotation of F1-ATPase visualized through angle-resolved single-fluorophore imaging.

Orientation dependence of single-fluorophore intensity was exploited in order to videotape conformational changes in a protein machine in real time. The fluorophore Cy3 attached to the central subunit of F(1)-ATPase revealed that the subunit rotates in the molecule in discrete 120 degrees steps and that each step is driven by the hydrolysis of one ATP molecule. These results, unlike those from the previous study under a frictional load, show that the 120 degrees stepping is a genuine property of this molecular motor. The data also show that the rate of ATP binding is insensitive to the load exerted on the rotor subunit.

[1]  Masasuke Yoshida,et al.  Catalytic Activity of the α3β3γ Complex of F1-ATPase without Noncatalytic Nucleotide Binding Site* , 1997, The Journal of Biological Chemistry.

[2]  Kazuhiko Kinosita,et al.  F1-ATPase Is a Highly Efficient Molecular Motor that Rotates with Discrete 120° Steps , 1998, Cell.

[3]  Analysis of time-dependent change of Escherichia coli F1-ATPase activity and its relationship with apparent negative cooperativity. , 1995, Biochimica et biophysica acta.

[4]  R. Aggeler,et al.  Rotation of a gamma-epsilon subunit domain in the Escherichia coli F1F0-ATP synthase complex. The gamma-epsilon subunits are essentially randomly distributed relative to the alpha3beta3delta domain in the intact complex. , 1997, The Journal of biological chemistry.

[5]  M. Saraste,et al.  FEBS Lett , 2000 .

[6]  P. Boyer.  The ATP synthase--a splendid molecular machine. , 1997, Annual review of biochemistry.

[7]  V. V. Bulygin,et al.  ATP hydrolysis by membrane-bound Escherichia coli F0F1 causes rotation of the gamma subunit relative to the beta subunits. , 1996, Biochimica et biophysica acta.

[8]  F. Oosawa,et al.  The loose coupling mechanism in molecular machines of living cells. , 1986, Advances in biophysics.

[9]  H. Noji,et al.  A rotary molecular motor that can work at near 100% efficiency. , 2000, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.

[10]  Kazuhiko Kinosita,et al.  Direct observation of the rotation of F1-ATPase , 1997, Nature.

[11]  K. Trybus,et al.  Myosin conformational states determined by single fluorophore polarization. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[12]  W. Junge,et al.  Three‐stepped rotation of subunits γ and ϵ in single molecules of F‐ATPase as revealed by polarized, confocal fluorometry , 1998 .

[13]  Jan Pieter Abrahams,et al.  Structure at 2.8 Â resolution of F1-ATPase from bovine heart mitochondria , 1994, Nature.

[14]  K. Kinosita.  Real time imaging of rotating molecular machines , 1999, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[15]  D. Seigneurin.  [Cytometry]. , 2020, Annales de Pathologie.

[16]  Tsuyoshi Hayakawa,et al.  Single-Molecule Detection by Laser-Induced Fluorescence Technique with a Position-Sensitive Photon-Counting Apparatus , 1994 .

[17]  W. Junge,et al.  Intersubunit rotation in active F-ATPase , 1996, Nature.

[18]  Kiwamu Saito,et al.  Imaging of single fluorescent molecules and individual ATP turnovers by single myosin molecules in aqueous solution , 1995, Nature.

[19]  I. Sase,et al.  Axial rotation of sliding actin filaments revealed by single-fluorophore imaging. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[20]  George Oster,et al.  Energy transduction in the F1 motor of ATP synthase , 1998, Nature.

[21]  W. Moerner,et al.  Those Blinking Single Molecules , 1997, Science.

[22]  T. Kodama,et al.  Kinetics of adenosine triphosphate hydrolysis by shortening myofibrils from rabbit psoas muscle. , 1991, The Journal of physiology.

[23]  A. Waggoner,et al.  Cyanine dye labeling reagents for sulfhydryl groups. , 1989, Cytometry.

[24]  S. Ishiwata,et al.  Dual-view microscopy with a single camera: real-time imaging of molecular orientations and calcium , 1991, The Journal of cell biology.

[25]  P. Boyer,et al.  The binding change mechanism for ATP synthase--some probabilities and possibilities. , 1993, Biochimica et biophysica acta.

[26]  D. Chemla,et al.  Single Molecule Dynamics Studied by Polarization Modulation. , 1996, Physical review letters.

[27]  T. Yanagida,et al.  Mechanochemical coupling in actomyosin energy transduction studied by in vitro movement assay. , 1990, Journal of molecular biology.

[28]  I. Sase,et al.  Real time imaging of single fluorophores on moving actin with an epifluorescence microscope. , 1995, Biophysical journal.

[29]  V. V. Bulygin,et al.  Rotation of subunits during catalysis by Escherichia coli F1-ATPase. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[30]  Adachi,et al.  Single‐fluorophore imaging with an unmodified epifluorescence microscope and conventional video camera , 1999, Journal of microscopy.