Guide to video recording of structure dynamics and dynamic processes of proteins by high-speed atomic force microscopy

High-speed atomic force microscopy (HS-AFM) allows direct visualization of dynamic structural changes and processes of functioning biological molecules in physiological solutions, at subsecond to sub-100-ms temporal and submolecular spatial resolution. Unlike fluorescence microscopy, wherein the subset of molecular events that you see is dependent on the site where the probe is placed, dynamic molecular events unselectively appear in detail in an AFM movie, facilitating our understanding of how biological molecules function. Here we present protocols for HS-AFM imaging of proteins in action, including preparation of cantilever tips, step-by-step procedures for HS-AFM imaging, and recycling of cantilevers and sample stages, together with precautions and troubleshooting advice for successful imaging. The protocols are adaptable in general for imaging many proteins and protein–nucleic acid complexes, and examples are described for looking at walking myosin, ATP-hydrolyzing rotorless F1-ATPase and cellulose-hydrolyzing cellulase. The entire protocol takes 10–15 h, depending mainly on the substrate surface to be used.

[1]  Toshio Ando,et al.  Video imaging of walking myosin V by high-speed atomic force microscopy , 2010, Nature.

[2]  Christopher Hein,et al.  High-speed atomic force microscopy reveals rotary catalysis of rotor-less F 1 -ATPase , 2011 .

[3]  Daniel J Müller,et al.  Atomic force microscopy and spectroscopy of native membrane proteins , 2007, Nature Protocols.

[4]  Hideki Kandori,et al.  High-speed atomic force microscopy shows dynamic molecular processes in photoactivated bacteriorhodopsin. , 2010, Nature nanotechnology.

[5]  Eric Lesniewska,et al.  Surface Topography of Membrane Domains , 2022 .

[6]  T. Ando,et al.  Traffic Jams Reduce Hydrolytic Efficiency of Cellulase on Cellulose Surface , 2011, Science.

[7]  Paul K. Hansma,et al.  Fast imaging and fast force spectroscopy of single biopolymers with a new atomic force microscope designed for small cantilevers , 1999 .

[8]  T. Ando,et al.  Identification of the single specific IQ motif of myosin V from which calmodulin dissociates in the presence of Ca2+. , 2006, Biochemistry.

[9]  T. Ando,et al.  Streptavidin 2D crystal substrates for visualizing biomolecular processes by atomic force microscopy. , 2009, Biophysical journal.

[10]  Toshio Ando,et al.  High-speed atomic force microscopy coming of age , 2012, Nanotechnology.

[11]  T. Ando,et al.  A high-speed atomic force microscope for studying biological macromolecules in action. , 2003, Chemphyschem : a European journal of chemical physics and physical chemistry.

[12]  T. Ando,et al.  Dynamic proportional-integral-differential controller for high-speed atomic force microscopy , 2006 .

[13]  Toshio Ando,et al.  Direct observation of surfactant aggregate behavior on a mica surface using high-speed atomic force microscopy. , 2011, Chemical communications.

[14]  Takeshi Okano,et al.  Flow properties of microcrystalline cellulose suspension prepared by acid treatment of native cellulose , 1998 .

[15]  J. Hoh,et al.  Calibration of optical lever sensitivity for atomic force microscopy , 1995 .

[16]  T. Ando,et al.  High-Speed AFM and Imaging of Biomolecular Processes , 2013 .

[17]  A. Turberfield,et al.  Direct observation of stepwise movement of a synthetic molecular transporter. , 2011, Nature nanotechnology.

[18]  Ami Chand,et al.  Probing protein–protein interactions in real time , 2000, Nature Structural Biology.

[19]  Takeshi Fukuma,et al.  High resonance frequency force microscope scanner using inertia balance support , 2008 .

[20]  Pierre Sens,et al.  Experimental evidence for membrane-mediated protein-protein interaction. , 2010, Biophysical journal.

[21]  Toshio Ando,et al.  High-speed atomic force microscopy techniques for observing dynamic biomolecular processes. , 2010, Methods in enzymology.

[22]  T. Ando,et al.  Anisotropic diffusion of point defects in a two-dimensional crystal of streptavidin observed by high-speed atomic force microscopy , 2008, Nanotechnology.

[23]  Toshio Ando,et al.  Feed-Forward Compensation for High-Speed Atomic Force Microscopy Imaging of Biomolecules , 2006 .

[24]  T. Ando,et al.  Direct observation of processive movement by individual myosin V molecules. , 2000, Biochemical and biophysical research communications.

[25]  T. Ando,et al.  Imaging of Nucleic Acids with Atomic Force Microscopy , 1990 .

[26]  T. Ando,et al.  Structural changes in bacteriorhodopsin in response to alternate illumination observed by high-speed atomic force microscopy. , 2011, Angewandte Chemie.

[27]  Toshio Ando,et al.  High-speed AFM and nano-visualization of biomolecular processes , 2008, Pflügers Archiv - European Journal of Physiology.

[28]  T. Ando,et al.  High-speed Atomic Force Microscopy for Capturing Dynamic Behavior of Protein Molecules at Work , 2005 .

[29]  Masatoshi Yokokawa,et al.  Fast‐scanning atomic force microscopy reveals the ATP/ADP‐dependent conformational changes of GroEL , 2006, The EMBO journal.

[30]  M. Wada,et al.  Surface density of cellobiohydrolase on crystalline celluloses , 2006, The FEBS journal.

[31]  T. Ando,et al.  Dynamics of nucleosomes assessed with time-lapse high-speed atomic force microscopy. , 2011, Biochemistry.

[32]  Toshio Ando,et al.  Fast phase imaging in liquids using a rapid scan atomic force microscope , 2006 .

[33]  R. Brasseur,et al.  Atomic force microscopy of supported lipid bilayers , 2008, Nature Protocols.

[34]  Hiroyuki Noji,et al.  High-Speed Atomic Force Microscopy Reveals Rotary Catalysis of Rotorless F1-ATPase , 2011, Science.

[35]  Toshio Ando,et al.  Tip-sample distance control using photothermal actuation of a small cantilever for high-speed atomic force microscopy. , 2007, The Review of scientific instruments.

[36]  Masashi Kitazawa,et al.  Batch Fabrication of Sharpened Silicon Nitride Tips , 2003 .

[37]  Toshio Ando,et al.  High-Speed Atomic Force Microscopy for Studying the Dynamic Behavior of Protein Molecules at Work , 2006 .

[38]  Cees Dekker,et al.  High-Speed AFM Reveals the Dynamics of Single Biomolecules at the Nanometer Scale , 2011, Cell.

[39]  C. le Grimellec,et al.  Deciphering the Structure, Growth and Assembly of Amyloid-Like Fibrils Using High-Speed Atomic Force Microscopy , 2010, PloS one.

[40]  T. Ando,et al.  High-speed atomic force microscopy for nano-visualization of dynamic biomolecular processes , 2008 .

[41]  T. Ando,et al.  Visualization of intrinsically disordered regions of proteins by high-speed atomic force microscopy. , 2008, Chemphyschem : a European journal of chemical physics and physical chemistry.

[42]  Toshio Ando,et al.  Active damping of the scanner for high-speed atomic force microscopy , 2005 .

[43]  F. Rico,et al.  High-speed atomic force microscopy: Structure and dynamics of single proteins. , 2011, Current opinion in chemical biology.

[44]  Y. Lyubchenko,et al.  Visual analysis of concerted cleavage by type IIF restriction enzyme SfiI in subsecond time region. , 2011, Biophysical journal.

[45]  J. Spudich,et al.  The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. , 1971, The Journal of biological chemistry.

[46]  C. Wyman,et al.  Protein-DNA interactions in high speed AFM: single molecule diffusion analysis of human RAD54. , 2011, Integrative biology : quantitative biosciences from nano to macro.

[47]  Toshio Ando,et al.  Dynamics of bacteriorhodopsin 2D crystal observed by high-speed atomic force microscopy. , 2009, Journal of structural biology.