---
OA_type: free access
_id: '6659'
abstract:
- lang: eng
  text: Chemical labeling of proteins with synthetic molecular probes offers the possibility
    to probe the functions of proteins of interest in living cells. However, the methods
    for covalently labeling targeted proteins using complementary peptide tag-probe
    pairs are still limited, irrespective of the versatility of such pairs in biological
    research. Herein, we report the new CysHis tag-Ni(II) probe pair for the specific
    covalent labeling of proteins. A broad-range evaluation of the reactivity profiles
    of the probe and the CysHis peptide tag afforded a tag-probe pair with an optimized
    and high labeling selectivity and reactivity. In particular, the labeling specificity
    of this pair was notably improved compared to the previously reported one. This
    pair was successfully utilized for the fluorescence imaging of membrane proteins
    on the surfaces of living cells, demonstrating its potential utility in biological
    research.
acknowledgement: his work was supported by the Grant-in-Aid for Scientific Research
  B (JSPS KAKENHI grant no. JP17H03090 to A. O.); the Scientific Research on Innovative
  Areas “Chemistry for Multimolecular Crowding Biosystems” (JSPS KAKENHI grant no.
  JP17H06349 to A. O.); and the European Union (European Research Council Advanced
  grant no. 694539 and Human Brain Project Ref. 720270 to R. S.). A. O. acknowledges
  the financial support of the Takeda Science Foundation.
article_processing_charge: No
article_type: original
author:
- first_name: Naoki
  full_name: Zenmyo, Naoki
  last_name: Zenmyo
- first_name: Hiroki
  full_name: Tokumaru, Hiroki
  last_name: Tokumaru
- first_name: Shohei
  full_name: Uchinomiya, Shohei
  last_name: Uchinomiya
- first_name: Hirokazu
  full_name: Fuchida, Hirokazu
  last_name: Fuchida
- first_name: Shigekazu
  full_name: Tabata, Shigekazu
  id: 4427179E-F248-11E8-B48F-1D18A9856A87
  last_name: Tabata
- first_name: Itaru
  full_name: Hamachi, Itaru
  last_name: Hamachi
- first_name: Ryuichi
  full_name: Shigemoto, Ryuichi
  id: 499F3ABC-F248-11E8-B48F-1D18A9856A87
  last_name: Shigemoto
  orcid: 0000-0001-8761-9444
- first_name: Akio
  full_name: Ojida, Akio
  last_name: Ojida
citation:
  ama: Zenmyo N, Tokumaru H, Uchinomiya S, et al. Optimized reaction pair of the CysHis
    tag and Ni(II)-NTA probe for highly selective chemical labeling of membrane proteins.
    <i>Bulletin of the Chemical Society of Japan</i>. 2019;92(5):995-1000. doi:<a
    href="https://doi.org/10.1246/bcsj.20190034">10.1246/bcsj.20190034</a>
  apa: Zenmyo, N., Tokumaru, H., Uchinomiya, S., Fuchida, H., Tabata, S., Hamachi,
    I., … Ojida, A. (2019). Optimized reaction pair of the CysHis tag and Ni(II)-NTA
    probe for highly selective chemical labeling of membrane proteins. <i>Bulletin
    of the Chemical Society of Japan</i>. Bulletin of the Chemical Society of Japan.
    <a href="https://doi.org/10.1246/bcsj.20190034">https://doi.org/10.1246/bcsj.20190034</a>
  chicago: Zenmyo, Naoki, Hiroki Tokumaru, Shohei Uchinomiya, Hirokazu Fuchida, Shigekazu
    Tabata, Itaru Hamachi, Ryuichi Shigemoto, and Akio Ojida. “Optimized Reaction
    Pair of the CysHis Tag and Ni(II)-NTA Probe for Highly Selective Chemical Labeling
    of Membrane Proteins.” <i>Bulletin of the Chemical Society of Japan</i>. Bulletin
    of the Chemical Society of Japan, 2019. <a href="https://doi.org/10.1246/bcsj.20190034">https://doi.org/10.1246/bcsj.20190034</a>.
  ieee: N. Zenmyo <i>et al.</i>, “Optimized reaction pair of the CysHis tag and Ni(II)-NTA
    probe for highly selective chemical labeling of membrane proteins,” <i>Bulletin
    of the Chemical Society of Japan</i>, vol. 92, no. 5. Bulletin of the Chemical
    Society of Japan, pp. 995–1000, 2019.
  ista: Zenmyo N, Tokumaru H, Uchinomiya S, Fuchida H, Tabata S, Hamachi I, Shigemoto
    R, Ojida A. 2019. Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe
    for highly selective chemical labeling of membrane proteins. Bulletin of the Chemical
    Society of Japan. 92(5), 995–1000.
  mla: Zenmyo, Naoki, et al. “Optimized Reaction Pair of the CysHis Tag and Ni(II)-NTA
    Probe for Highly Selective Chemical Labeling of Membrane Proteins.” <i>Bulletin
    of the Chemical Society of Japan</i>, vol. 92, no. 5, Bulletin of the Chemical
    Society of Japan, 2019, pp. 995–1000, doi:<a href="https://doi.org/10.1246/bcsj.20190034">10.1246/bcsj.20190034</a>.
  short: N. Zenmyo, H. Tokumaru, S. Uchinomiya, H. Fuchida, S. Tabata, I. Hamachi,
    R. Shigemoto, A. Ojida, Bulletin of the Chemical Society of Japan 92 (2019) 995–1000.
date_created: 2019-07-21T21:59:16Z
date_published: 2019-05-15T00:00:00Z
date_updated: 2025-09-10T10:37:21Z
day: '15'
ddc:
- '570'
department:
- _id: RySh
doi: 10.1246/bcsj.20190034
ec_funded: 1
external_id:
  isi:
  - '000468101800007'
has_accepted_license: '1'
intvolume: '        92'
isi: 1
issue: '5'
language:
- iso: eng
main_file_link:
- open_access: '1'
  url: https://doi.org/10.1246/bcsj.20190034
month: '05'
oa: 1
oa_version: Published Version
page: 995-1000
project:
- _id: 25CA28EA-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '694539'
  name: 'In situ analysis of single channel subunit composition in neurons: physiological
    implication in synaptic plasticity and behaviour'
publication: Bulletin of the Chemical Society of Japan
publication_identifier:
  issn:
  - 0009-2673
publication_status: published
publisher: Bulletin of the Chemical Society of Japan
quality_controlled: '1'
scopus_import: '1'
status: public
title: Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe for highly selective
  chemical labeling of membrane proteins
type: journal_article
user_id: 317138e5-6ab7-11ef-aa6d-ffef3953e345
volume: 92
year: '2019'
...
---
_id: '7391'
abstract:
- lang: eng
  text: 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.
article_processing_charge: No
article_type: original
author:
- first_name: Shigekazu
  full_name: Tabata, Shigekazu
  id: 4427179E-F248-11E8-B48F-1D18A9856A87
  last_name: Tabata
- first_name: Marijo
  full_name: Jevtic, Marijo
  id: 4BE3BC94-F248-11E8-B48F-1D18A9856A87
  last_name: Jevtic
- first_name: Nobutaka
  full_name: Kurashige, Nobutaka
  last_name: Kurashige
- first_name: Hirokazu
  full_name: Fuchida, Hirokazu
  last_name: Fuchida
- first_name: Munetsugu
  full_name: Kido, Munetsugu
  last_name: Kido
- first_name: Kazushi
  full_name: Tani, Kazushi
  last_name: Tani
- first_name: Naoki
  full_name: Zenmyo, Naoki
  last_name: Zenmyo
- first_name: Shohei
  full_name: Uchinomiya, Shohei
  last_name: Uchinomiya
- first_name: Harumi
  full_name: Harada, Harumi
  id: 2E55CDF2-F248-11E8-B48F-1D18A9856A87
  last_name: Harada
  orcid: 0000-0001-7429-7896
- first_name: Makoto
  full_name: Itakura, Makoto
  last_name: Itakura
- first_name: Itaru
  full_name: Hamachi, Itaru
  last_name: Hamachi
- first_name: Ryuichi
  full_name: Shigemoto, Ryuichi
  id: 499F3ABC-F248-11E8-B48F-1D18A9856A87
  last_name: Shigemoto
  orcid: 0000-0001-8761-9444
- first_name: Akio
  full_name: Ojida, Akio
  last_name: Ojida
citation:
  ama: Tabata S, Jevtic M, Kurashige N, et al. Electron microscopic detection of single
    membrane proteins by a specific chemical labeling. <i>iScience</i>. 2019;22(12):256-268.
    doi:<a href="https://doi.org/10.1016/j.isci.2019.11.025">10.1016/j.isci.2019.11.025</a>
  apa: 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. <i>IScience</i>. Elsevier. <a href="https://doi.org/10.1016/j.isci.2019.11.025">https://doi.org/10.1016/j.isci.2019.11.025</a>
  chicago: 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.” <i>IScience</i>.
    Elsevier, 2019. <a href="https://doi.org/10.1016/j.isci.2019.11.025">https://doi.org/10.1016/j.isci.2019.11.025</a>.
  ieee: S. Tabata <i>et al.</i>, “Electron microscopic detection of single membrane
    proteins by a specific chemical labeling,” <i>iScience</i>, vol. 22, no. 12. Elsevier,
    pp. 256–268, 2019.
  ista: 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.
  mla: Tabata, Shigekazu, et al. “Electron Microscopic Detection of Single Membrane
    Proteins by a Specific Chemical Labeling.” <i>IScience</i>, vol. 22, no. 12, Elsevier,
    2019, pp. 256–68, doi:<a href="https://doi.org/10.1016/j.isci.2019.11.025">10.1016/j.isci.2019.11.025</a>.
  short: 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.
corr_author: '1'
date_created: 2020-01-29T15:56:56Z
date_published: 2019-12-20T00:00:00Z
date_updated: 2026-07-08T22:30:06Z
day: '20'
ddc:
- '570'
department:
- _id: RySh
doi: 10.1016/j.isci.2019.11.025
ec_funded: 1
external_id:
  isi:
  - '000504652000020'
  pmid:
  - '31786521'
file:
- access_level: open_access
  checksum: f3e90056a49f09b205b1c4f8c739ffd1
  content_type: application/pdf
  creator: dernst
  date_created: 2020-02-04T10:48:36Z
  date_updated: 2020-07-14T12:47:57Z
  file_id: '7448'
  file_name: 2019_iScience_Tabata.pdf
  file_size: 7197776
  relation: main_file
file_date_updated: 2020-07-14T12:47:57Z
has_accepted_license: '1'
intvolume: '        22'
isi: 1
issue: '12'
language:
- iso: eng
month: '12'
oa: 1
oa_version: Published Version
page: 256-268
pmid: 1
project:
- _id: 25CA28EA-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '694539'
  name: 'In situ analysis of single channel subunit composition in neurons: physiological
    implication in synaptic plasticity and behaviour'
- _id: 25CBA828-B435-11E9-9278-68D0E5697425
  call_identifier: H2020
  grant_number: '720270'
  name: Human Brain Project Specific Grant Agreement 1
publication: iScience
publication_identifier:
  issn:
  - 2589-0042
publication_status: published
publisher: Elsevier
quality_controlled: '1'
related_material:
  record:
  - id: '11393'
    relation: dissertation_contains
    status: public
scopus_import: '1'
status: public
title: Electron microscopic detection of single membrane proteins by a specific chemical
  labeling
tmp:
  image: /images/cc_by.png
  legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
  name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
  short: CC BY (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 22
year: '2019'
...