A biotin-HaloTag ligand enables efficient affinity capture of
protein variants from live cells

J 2025

A biotin-HaloTag ligand enables efficient affinity capture of protein variants from live cells

YADAV, Anoop Kumar; Abhijeet Sahebrao JADHAV; Pawel Marcin SZCZEPANIK; Paolo FAGHERAZZI; Ivo KABELKA et al.

Základní údaje

Originální název

A biotin-HaloTag ligand enables efficient affinity capture of protein variants from live cells

Autoři

YADAV, Anoop Kumar; Abhijeet Sahebrao JADHAV; Pawel Marcin SZCZEPANIK; Paolo FAGHERAZZI; Ivo KABELKA ; Robert VÁCHA; Jakub ŠVENDA a Hana POLASEK-SEDLACKOVA

Vydání

JOURNAL OF CELL BIOLOGY, NEW YORK, ROCKEFELLER UNIV PRESS, 2025, 0021-9525

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10601 Cell biology

Stát vydavatele

Spojené státy

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 6.400 v roce 2024

Označené pro přenos do RIV

Ano

Kód RIV

RIV/00216224:14310/25:00142954

Organizační jednotka

Přírodovědecká fakulta

Klíčová slova anglicky

GENERAL-METHOD; FLUOROGENIC PROBES

Štítky

14313010, 14314010, rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 17. 12. 2025 13:16, Mgr. Pavla Foltynová, Ph.D.

Anotace

V originále

HaloTag technology represents a versatile tool for studying proteins. Fluorescent HaloTag ligands employed in sequential labeling led to the discovery of distinct protein variants for histones, cohesins, and MCM complexes. However, an efficient biochemical approach to separate these distinct protein variants to study their biological functions is missing. Principally, being a gap in technology, the HaloTag toolbox lacks affinity ligands displaying good cell permeability and efficient affinity capture. Here, we describe the design, synthesis, and validation of a new cell-permeable biotin-HaloTag ligand, which allows rapid labeling of Halo-tagged proteins in live cells and their efficient separation using streptavidin pull-down. We provide a proof-of-concept application of how to use the herein-developed affinity ligand in sequential labeling to biochemically separate protein variants and study their biological properties. This approach enables to address fundamental questions concerning essential cellular processes, including genome duplication and chromatin maintenance.

Návaznosti

LM2023052, projekt VaV

Název: Národní infrastruktura chemické biologie

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, Národní infrastruktura chemické biologie

LX22NPO5103, projekt VaV

Název: Národní institut virologie a bakteriologie (Akronym: NIVB)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, Národní institut virologie a bakteriologie, 5.1 EXCELES

Citovat

YADAV, Anoop Kumar; Abhijeet Sahebrao JADHAV; Pawel Marcin SZCZEPANIK; Paolo FAGHERAZZI; Ivo KABELKA; Robert VÁCHA; Jakub ŠVENDA a Hana POLASEK-SEDLACKOVA. A biotin-HaloTag ligand enables efficient affinity capture of protein variants from live cells. JOURNAL OF CELL BIOLOGY. NEW YORK: ROCKEFELLER UNIV PRESS, 2025, roč. 224, č. 8, s. 1-26. ISSN 0021-9525. Dostupné z: https://doi.org/10.1083/jcb.202410025.

@article{2541259,
   author = {Yadav, Anoop Kumar and Jadhav, Abhijeet Sahebrao and Szczepanik, Pawel Marcin and Fagherazzi, Paolo and Kabelka, Ivo and Vácha, Robert and Švenda, Jakub and PolasekandSedlackova, Hana},
   article_location = {NEW YORK},
   article_number = {8},
   doi = {https://doi.org/10.1083/jcb.202410025},
   keywords = {GENERAL-METHOD; FLUOROGENIC PROBES},
   language = {eng},
   issn = {0021-9525},
   journal = {JOURNAL OF CELL BIOLOGY},
   title = {A biotin-HaloTag ligand enables efficient affinity capture of protein variants from live cells},
   url = {https://rupress.org/jcb/article-abstract/224/8/e202410025/278122/A-biotin-HaloTag-ligand-enables-efficient-affinity?redirectedFrom=fulltext},
   volume = {224},
   year = {2025}
}

TY  - JOUR
ID  - 2541259
AU  - Yadav, Anoop Kumar - Jadhav, Abhijeet Sahebrao - Szczepanik, Pawel Marcin - Fagherazzi, Paolo - Kabelka, Ivo - Vácha, Robert - Švenda, Jakub - Polasek-Sedlackova, Hana
PY  - 2025
TI  - A biotin-HaloTag ligand enables efficient affinity capture of protein variants from live cells
JF  - JOURNAL OF CELL BIOLOGY
VL  - 224
IS  - 8
SP  - 1-26
EP  - 1-26
PB  - ROCKEFELLER UNIV PRESS
SN  - 00219525
KW  - GENERAL-METHOD
KW  - FLUOROGENIC PROBES
UR  - https://rupress.org/jcb/article-abstract/224/8/e202410025/278122/A-biotin-HaloTag-ligand-enables-efficient-affinity?redirectedFrom=fulltext
N2  - HaloTag technology represents a versatile tool for studying proteins. Fluorescent HaloTag ligands employed in sequential labeling led to the discovery of distinct protein variants for histones, cohesins, and MCM complexes. However, an efficient biochemical approach to separate these distinct protein variants to study their biological functions is missing. Principally, being a gap in technology, the HaloTag toolbox lacks affinity ligands displaying good cell permeability and efficient affinity capture. Here, we describe the design, synthesis, and validation of a new cell-permeable biotin-HaloTag ligand, which allows rapid labeling of Halo-tagged proteins in live cells and their efficient separation using streptavidin pull-down. We provide a proof-of-concept application of how to use the herein-developed affinity ligand in sequential labeling to biochemically separate protein variants and study their biological properties. This approach enables to address fundamental questions concerning essential cellular processes, including genome duplication and chromatin maintenance.
ER  -

YADAV, Anoop Kumar; Abhijeet Sahebrao JADHAV; Pawel Marcin SZCZEPANIK; Paolo FAGHERAZZI; Ivo KABELKA; Robert VÁCHA; Jakub ŠVENDA a Hana POLASEK-SEDLACKOVA. A biotin-HaloTag ligand enables efficient affinity capture of protein variants from live cells. \textit{JOURNAL OF CELL BIOLOGY}. NEW YORK: ROCKEFELLER UNIV PRESS, 2025, roč.~224, č.~8, s.~1-26. ISSN~0021-9525. Dostupné z: https://doi.org/10.1083/jcb.202410025.

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