However, removal of captured proteins from Streptavidin can be problematic due to the strong nature of the binding. One possible solution to this problem are linkers that allow for a selective cleavage following purification.
Biotinylation of an azide-bearing glycoprotein using Click-chemistry, followed by purification of the labelled protein over Streptavidin beads, release of the protein by hydrazinolysis of the Dde-group, and analysis of the isolated glycoprotein (adapted from Griffin et al. Mol. Biosys. 2016).
We recently introduced three bifunctional clickable linkers that each bear a biotin moiety on one end as well as a clickable group (azide or alkyne) on the other. Those two ends are connected by a Dde-based linker. Dde [N-1-(4,4-dimethyl-2,6-dioxocyclohexylidene)ethyl] is stable to denaturing wash conditions as well as acidic and basic conditions, while allowing for a mild and selective cleavage with a buffered aqueous solution of 2% hydrazine. Consequently, our new bifunctional linkers can be selectively attached to appropriately modified biomolecules via
Click-chemistry, allowing for subsequent purification on Streptavidin beads followed by mild release of the captured protein from the beads.
→ Order our catalogue Biotinylation Reagents, or download the E-Book, to get a comprehensive overview of our compounds for biotinylation.
- Mapping the Binding Site of BMS-708163 on gamma-Secretase with Cleavable Photoprobes; N. Gertsik, C. W. Am Ende, K. F. Geoghegan, C. Nguyen, P. Mukherjee, S. Mente, U. Seneviratne, D. S. Johnson and Y. M. Li; Cell Chem Biol 2017; 24: 3-8. doi:10.1016/j.chembiol.2016.12.006
- Comprehensive mapping of O-GlcNAc modification sites using a chemically cleavable tag; M. E. Griffin, E. H. Jensen, D. E. Mason, C. L. Jenkins, S. E. Stone, E. C. Peters and L. C. Hsieh-Wilson; Mol Biosyst 2016; 12: 1756-9. doi:10.1039/c6mb00138f
- Cleavable trifunctional biotin reagents for protein labelling, capture and release; Y. Yang and S. H. Verhelst; Chem Commun 2013; 49: 5366-8. doi:10.1039/c3cc42076k