Abstract
A major challenge in attaching fluorophores or other handles to proteins is the availability of a site-specific labeling strategy that provides stoichiometric modification without compromising protein integrity. We developed a simple approach that combines TEV protease cleavage, sortase modification and affinity purification to N-terminally label proteins. To achieve stoichiometrically-labeled protein, we included a short affinity tag in the fluorophore-containing peptide for post-labeling purification of the modified protein. This strategy can be easily applied to any recombinant protein with a TEV site and we demonstrate this on Epidermal Growth Factor Receptor (EGFR) and Membrane Scaffold Protein (MSP) constructs.
Original language | English |
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Pages (from-to) | 55-58 |
Number of pages | 4 |
Journal | Analytical Biochemistry |
Volume | 521 |
DOIs | |
State | Published - Mar 15 2017 |
Keywords
- Affinity tag
- LPXTG
- Post-labeling purification
- Sortase
- TEV