Electron microscopic detection of single membrane proteins by a specific chemical labeling

S. Tabata, M. Jevtic, N. Kurashige, H. Fuchida, M. Kido, K. Tani, N. Zenmyo, S. Uchinomiya, H. Harada, M. Itakura, I. Hamachi, R. Shigemoto, A. Ojida, IScience 22 (2019) 256–268.

Download
OA 2019_iScience_Tabata.pdf 7.20 MB

Journal Article | Published | English
Author
Tabata, ShigekazuIST Austria; Jevtic, MarijoIST Austria; Kurashige, Nobutaka; Fuchida, Hirokazu; Kido, Munetsugu; Tani, Kazushi; Zenmyo, Naoki; Uchinomiya, Shohei; Harada, HarumiIST Austria ; Itakura, Makoto; Hamachi, Itaru; Shigemoto, RyuichiIST Austria
All
Department
Abstract
Electron microscopy (EM) is a technology that enables visualization of single proteins at a nanometer resolution. However, current protein analysis by EM mainly relies on immunolabeling with gold-particle-conjugated antibodies, which is compromised by large size of antibody, precluding precise detection of protein location in biological samples. Here, we develop a specific chemical labeling method for EM detection of proteins at single-molecular level. Rational design of α-helical peptide tag and probe structure provided a complementary reaction pair that enabled specific cysteine conjugation of the tag. The developed chemical labeling with gold-nanoparticle-conjugated probe showed significantly higher labeling efficiency and detectability of high-density clusters of tag-fused G protein-coupled receptors in freeze-fracture replicas compared with immunogold labeling. Furthermore, in ultrathin sections, the spatial resolution of the chemical labeling was significantly higher than that of antibody-mediated labeling. These results demonstrate substantial advantages of the chemical labeling approach for single protein visualization by EM.
Publishing Year
Date Published
2019-12-20
Journal Title
iScience
Volume
22
Issue
12
Page
256-268
ISSN
IST-REx-ID

Cite this

Tabata S, Jevtic M, Kurashige N, et al. Electron microscopic detection of single membrane proteins by a specific chemical labeling. iScience. 2019;22(12):256-268. doi:10.1016/j.isci.2019.11.025
Tabata, S., Jevtic, M., Kurashige, N., Fuchida, H., Kido, M., Tani, K., … Ojida, A. (2019). Electron microscopic detection of single membrane proteins by a specific chemical labeling. IScience, 22(12), 256–268. https://doi.org/10.1016/j.isci.2019.11.025
Tabata, Shigekazu, Marijo Jevtic, Nobutaka Kurashige, Hirokazu Fuchida, Munetsugu Kido, Kazushi Tani, Naoki Zenmyo, et al. “Electron Microscopic Detection of Single Membrane Proteins by a Specific Chemical Labeling.” IScience 22, no. 12 (2019): 256–68. https://doi.org/10.1016/j.isci.2019.11.025.
S. Tabata et al., “Electron microscopic detection of single membrane proteins by a specific chemical labeling,” iScience, vol. 22, no. 12, pp. 256–268, 2019.
Tabata S, Jevtic M, Kurashige N, Fuchida H, Kido M, Tani K, Zenmyo N, Uchinomiya S, Harada H, Itakura M, Hamachi I, Shigemoto R, Ojida A. 2019. Electron microscopic detection of single membrane proteins by a specific chemical labeling. iScience. 22(12), 256–268.
Tabata, Shigekazu, et al. “Electron Microscopic Detection of Single Membrane Proteins by a Specific Chemical Labeling.” IScience, vol. 22, no. 12, Elsevier, 2019, pp. 256–68, doi:10.1016/j.isci.2019.11.025.
All files available under the following license(s):
Creative Commons Attribution 4.0 International Public License (CC-BY 4.0):
Main File(s)
File Name
Access Level
OA Open Access
Date Uploaded
2020-02-04
MD5 Checksum
f3e90056a49f09b205b1c4f8c739ffd1


Export

Marked Publications

Open Data IST Research Explorer

Sources

PMID: 31786521
PubMed | Europe PMC

Search this title in

Google Scholar