Binding Affinity Measurement of Nuclear Export Signal Peptides to Their Exporter CRM1.
Binding affinity
CRM1
Fluorescence polarization
NES
Nuclear export signals
XPO1
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:
2022
2022
Historique:
entrez:
12
4
2022
pubmed:
13
4
2022
medline:
15
4
2022
Statut:
ppublish
Résumé
CRM1 recognizes hundreds to thousands of protein cargoes by binding to the eight to fifteen residue-long nuclear export signals (NESs) within their polypeptide chains. Various assays to measure the binding affinity of NESs for CRM1 have been developed. CRM1 binds to NESs with a wide range of binding affinities, with dissociation constants that span from low nanomolar to tens of micromolar. An optimized binding affinity assay with improved throughput was recently developed to measure binding affinities of NES peptides for CRM1 in the presence of excess RanGTP. The assay can measure affinities, with multiple replicates, for up to seven different NES peptides per screening plate. Here, we present a protocol for the purification of the necessary proteins and for measuring CRM1-NES binding affinities.
Identifiants
pubmed: 35412243
doi: 10.1007/978-1-0716-2337-4_16
doi:
Substances chimiques
Karyopherins
0
Nuclear Export Signals
0
Peptides
0
Receptors, Cytoplasmic and Nuclear
0
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
245-256Subventions
Organisme : NIGMS NIH HHS
ID : R01 GM069909
Pays : United States
Informations de copyright
© 2022. Springer Science+Business Media, LLC, part of Springer Nature.
Références
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