Poly-Ubiquitin chains are composed of Ubiquitin monomers that are covalently linked through  isopeptide bonds, which typically form between a lysine residue of one Ubiquitin molecule and the C-terminal glycine residue of another Ubiquitin molecule (1). Each human Ubiquitin monomer is 76 amino acids (aa) in length and shares 96% and 100% aa identity with yeast and mouse Ubiquitin, respectively (2). Seven of the 76 aa in Ubiquitin are lysine residues that can participate in poly-Ubiquitin chain formation. Linkage through specific lysine residues is thought to serve as a signal that affects protein degradation, signaling, trafficking, and other cellular processes (3-8).

This mixture of poly-Ubiquitin chains contains di-Ubiquitin and higher MW species; mono-Ubiquitin has been removed. These chains have been modified with biotin via primary amine coupling.  This results in multiple biotinylated species modified at the N-terminus, as well as lysine residues.  Biotinylated Ubiquitin can be detected using avidin-linked reagents.