LC-MS of N-terminal Biotin-Ubiquitin.
A single biotin moiety is conjugated on the N-terminus of ubiquitin. All lysine residues of ubiquitin are still available for formation of polyubiquitin chains. The extremely high binding affinity between biotin and streptavidin allows the use of N-terminal biotin-ubiquitin for the following in vitro assays: 1) rapid and efficient purification of polyubiquitinated substrate proteins; 2) monitoring substrate ubiquitination using an HRP-conjugated streptavidin antibody; and 3) setting up high throughput FRET assays to monitor substrate ubiquitination in assays screening E3 ubiquitin ligase inhibitors.
