In Vivo Histone Labeling Using Ultrafast trans-Splicing Inteins.
Blotting, Western
/ methods
Cell Fractionation
/ methods
Cell-Penetrating Peptides
/ chemistry
Chromatin
/ chemistry
Chromatography, High Pressure Liquid
/ methods
Chromatography, Reverse-Phase
/ methods
Disulfides
/ chemistry
Fluorescent Dyes
/ chemistry
Gene Expression
HEK293 Cells
Histones
/ chemistry
Humans
Inteins
Microscopy, Confocal
Nuclear Proteins
/ metabolism
Protein Splicing
Recombinant Proteins
/ chemistry
Solid-Phase Synthesis Techniques
/ methods
Synthetic Biology
/ methods
Transfection
Chromatin
In cellulo labeling
Split inteins
Synthetic biology
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2020
2020
Historique:
entrez:
8
3
2020
pubmed:
8
3
2020
medline:
6
3
2021
Statut:
ppublish
Résumé
The development of expressed protein ligation (EPL) widened the scope of questions that could be addressed by mechanistic biochemistry. Protein trans-splicing (PTS) relies on the same basic chemical principles, but utilizes split inteins to tracelessly ligate distinct peptide or polypeptide fragments together with native peptide bonds. Here we present a method to adapt PTS methodologies for their use in live cells, in order to deliver synthetic or native histone modifications. As an example, we provide a protocol to incorporate a small molecule fluorophore into chromatinized histones. The protocol should be easily adaptable to incorporate other modifications to chromatin in vivo.
Identifiants
pubmed: 32144669
doi: 10.1007/978-1-0716-0434-2_10
pmc: PMC7534146
mid: NIHMS1632573
doi:
Substances chimiques
Cell-Penetrating Peptides
0
Chromatin
0
Disulfides
0
Fluorescent Dyes
0
Histones
0
Nuclear Proteins
0
Recombinant Proteins
0
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Langues
eng
Sous-ensembles de citation
IM
Pagination
201-219Subventions
Organisme : NCI NIH HHS
ID : P30 CA008748
Pays : United States
Organisme : NCI NIH HHS
ID : P50 CA192937
Pays : United States
Organisme : NIDA NIH HHS
ID : R21 DA044767
Pays : United States
Organisme : NIGMS NIH HHS
ID : T32 GM115327
Pays : United States
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