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. 2022 Mar 5;12(5):e4344.
doi: 10.21769/BioProtoc.4344.

High-speed Atomic Force Microscopy Observation of Internal Structure Movements in Living Mycoplasma

Kohei Kobayashi  1 , Noriyuki Kodera  2 , Makoto Miyata  1   3
Affiliations

High-speed Atomic Force Microscopy Observation of Internal Structure Movements in Living Mycoplasma

Kohei Kobayashi et al. Bio Protoc. .

Abstract

Dozens of Mycoplasma species belonging to the class Mollicutes bind to solid surfaces through the organelle formed at a cell pole and glide in its direction by a unique mechanism. In Mycoplasma mobile, the fastest gliding species in Mycoplasma, the force for gliding is generated by ATP hydrolysis on an internal structure. However, the spatial and temporal behaviors of the internal structures in living cells were unclear. High-speed atomic force microscopy (HS-AFM) is a powerful method to monitor the dynamic behaviors of biomolecules and cells that can be captured while maintaining their active state in aqueous solution. In this protocol, we describe a method to detect their movements using HS-AFM. This protocol should be useful for the studies of many kinds of microorganisms. Graphic abstract: Scanning Mycoplasma cell.

Keywords: Class Mollicutes; AFM; ATPase; Internal structure; Pathogenic bacteria; Probing.

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Conflict of interest statement

Competing interestsThere are no conflicts of interest or competing interests.

Figures

Figure 1.
Figure 1.. HS-AFM scanner with glass stage and sample drop.
Piezoelectric elements are used to drive each axis in the XYZ directions. Z piezo, glass stage, and sample solution are seen in a pink dashed circle, as illustrated in the lower right. The glass stage is fixed onto the Z-piezo using nail polish.
Figure 2.
Figure 2.. Procedure for liquid replacement on the glass stage.
Figure 3.
Figure 3.. HS-AFM.
(a) Picture showing the mechanical parts and some of the electrical devices of HS-AFM. The AFM unit (scanner and cantilever) is mounted on an inverted optical microscope which is placed on an anti-vibration table. (b) Schematic of experimental setup. Cantilever and M. mobile cells are not in scale. The optical view around the cantilever and the glass substrate can be observed through the monitor.
Figure 4.
Figure 4.. Representative images taken by HS-AFM.
(a) Whole cell image (b) Particle structure image [Magnified image of the boxed area of the panel (a)]. This figure was modified a previous paper ( Kobayashi et al., 2021 ).
See this image and copyright information in PMC

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References

    1. Ando T.(2018). High-speed atomic force microscopy and its future prospects. Biophys Rev 10(2): 285-292. - PMC - PubMed
    1. Kobayashi K., Kodera N., Kasai T., Tahara Y. O., Toyonaga T., Mizutani M., Fujiwara I., Ando T. and Miyata M.(2021). Movements of Mycoplasma mobile gliding machinery detected by high-speed atomic force microscopy . mBio 12(3): e0004021. - PMC - PubMed
    1. Kodera N. and Ando T.(2020). High-speed atomic force microscopy to study myosin motility. Adv Exp Med Biol 1239: 127-152. - PubMed
    1. Kodera N., Noshiro D., Dora S. K., Mori T., Habchi J., Blocquel D., Gruet A., Dosnon M., Salladini E., Bignon C., et al. .(2021). Structural and dynamics analysis of intrinsically disordered proteins by high-speed atomic force microscopy. Nat Nanotechnol 16(2): 181-189. - PubMed
    1. Kodera N., Yamamoto D., Ishikawa R. and Ando T.(2010). Video imaging of walking myosin V by high-speed atomic force microscopy. Nature 468(7320): 72-76. - PubMed