Chromophore pre-maturation for improved speed and sensitivity of split-GFP monitoring of protein secretion.
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
22 01 2019
22 01 2019
Historique:
received:
01
03
2018
accepted:
22
11
2018
entrez:
24
1
2019
pubmed:
24
1
2019
medline:
23
6
2020
Statut:
epublish
Résumé
Complementation-dependent fluorescence is a powerful way to study co-localization or interactions between biomolecules. A split-GFP variant, involving the self-associating GFP 1-10 and GFP 11, has previously provided a convenient approach to measure recombinant protein titers in cell supernatants. A limitation of this approach is the slow chromophore formation after complementation. Here, we alleviate this lag in signal generation by allowing the GFP 1-10 chromophore to mature on a solid support containing GFP 11 before applying GFP 1-10 in analyses. The pre-maturated GFP 1-10 provided up to 150-fold faster signal generation compared to the non-maturated version. Moreover, pre-maturated GFP 1-10 significantly improved the ability of discriminating between Chinese hamster ovary (CHO) cell lines secreting GFP 11-tagged erythropoietin protein at varying rates. Its improved kinetics make the pre-maturated GFP 1-10 a suitable reporter molecule for cell biology research in general, especially for ranking individual cell lines based on secretion rates of recombinant proteins.
Identifiants
pubmed: 30670736
doi: 10.1038/s41598-018-36559-x
pii: 10.1038/s41598-018-36559-x
pmc: PMC6342966
doi:
Substances chimiques
GFP10 Protein
0
Luminescent Proteins
0
Proteins
0
Recombinant Proteins
0
Erythropoietin
11096-26-7
Green Fluorescent Proteins
147336-22-9
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
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