High-speed atomic force microscopy and its future prospects

Abstract

Various techniques have been developed and used to investigate how proteins produce complex biological architectures and phenomena. Among these techniques, high-speed atomic force microscopy (HS-AFM) holds a unique position. It is only HS-AFM that allows the simultaneous assessment of structure and dynamics of single protein molecules in action. This new microscopy tool has been successfully applied to a variety of proteins, from motor proteins to membrane proteins, antibodies, enzymes, and even to intrinsically disordered proteins. And yet there still remain many biomolecular phenomena that cannot be addressed by HS-AFM in its current form. Here, I present a brief history of HS-AFM development, describe the current state of HS-AFM, and then discuss which new biological scanning probe microscopy techniques will be coming up next.

Keywords: Dynamic processes; High-speed AFM; Imaging; Proteins; Structural changes.

Fig. 1

High-speed atomic force microscopy (HS-AFM) imaging of the outer surface of the live magnetotactic spirillum bacterium Magnetospirillum magneticum AMB-1. a Low-magnification image of the outer surface. b High-magnification image of the outer surface. c Trajectories of the center position of four indents surrounded by hexagons. These trajectories were drawn by tracing the movement of each indent. d HS-AFM images showing dissociation process of a reticulate structure in an outer membrane patch. Numbers indicate frame number. e Highly magnified AFM image of molecules observed after dissociation. f Sodium dodecyl sulfate–polyacrylamide gel electrophoresis gel profile of extracted proteins from the isolated outer membrane (lane 2). The protein band indicated by the arrow was identified as porin. The molecular masses of the standards are indicated on the left sides of the lanes

Fig. 2

HS-AFM images of mitochondria. a Magnified image showing assembled voltage-dependent anionic channels. b,c HS-AFM images before (b) and after (c) rupture of a mitochondrion