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Ubiquitin and Ubiquitin-like Modifiers

Introduction of ubiquitination

The ubiquitin super family is characterized by small protein modifiers that are covalently attached to proteins substrates through a series of biochemically similar steps (Figure 1).

Figure 1. Schematic of the general cascade for modification by ubiquitin and ubiquitin-like (Ubl) modifiers.

E1, E2, and E3 are the activating enzyme, conjugating enzyme, and ligase, respectively for the various Ubl pathways; the number of distinct enzymes at each step varies for the particular modifier system. M represents ubiquitin or any other Ubl. S is the substrate and D is the demodifying enzyme that removes the modifier and returns the substrate to the unmodified form.

The first step in the conjugation process is activation of the modifier through an ATP-dependent reaction that forms a linkage between the modifier and the activation enzyme (E1). Subsequently the modifier is transferred to the conjugation enzyme (E2), through a thioester linkage. Finally, the modifier is transferred to the substrate using an E3 ligase that provides substrate specificity. This final conjugation of the modifier to the substrate occurs through an isopeptide bond linking the C-terminus of the modifier with the epsilon amino group of a lysine residue in the target protein. Depending on the substrate, more than one lysine may be modified and more than one type of modifier may be utilized; in some cases the different modifications occur on separate lysine residues while in other cases different modifiers compete for the same lysine, often with opposing functional effects. Additionally, depending on the modifier used both mono- and poly-chains of the modifier can be formed resulting in different outcomes for the substrate.


The prototype of this superfamily is ubiquitin, an 8-9 kDa protein present in all cells whose crystal structure shows a distinctive fold dominated by a β-sheet with five antiparallel β -strands and a single helical segment. Some studies have shown that eukaryotes possess a distinctive enzymatic apparatus for Ubiquitin-modification, comprised of a cascade of three enzymes, E1, E2, and E3. Ubiquitin is attached to proteins by the sequential action of E1-, E2- and E3-ligase proteins. These enzymes successively activate  ubiquitin for transfer using the free energy derived from ATP hydrolysis, relay it via thiocarboxylate linkages involving the C-terminal residue of ubiquitin or ubiquitin-like proteins, and finally transfer it to the epsilon NH 2 group on lysines, the amino terminal NH 2 groups, or on rare occasions cysteines on target polypeptides. E3-ligases contain either a HECT (homologous to E6-AP carboxy terminus) or RING (really interesting new gene) finger domain, a common feature of many different proteins, which catalyzes the transfer of the ubiquitin from the E2 to a Lys residue on the substrate protein. Small RING domain containing proteins form multimeric complexes, such as the Skp1-Cullin-F-box protein, whereas larger RING domain containing proteins can function as a single subunit. The specificity of the ubiquitination reaction is regulated by the E3-ligases. Recently identified E4-ligases may promote ubiquitin chain formation, poly-ubiquitination.

In early times, ubiquitin was believed to mainly serve as a tag for protein degradation, and it is now known that after being coupled to thousands of proteins, either as monomer or as chains, ubiquitin can serve various purposes. More than 200 proteins harbor one or more copies of the around 20 Ubiquitin-binding domains (UBDs) that recognize specific inter-Ubiquitin linkages and mediate transient Ubiquitin–UBD interactions. These interactions, combined with dynamic ubiquitination/deubiquitination reactions, create flexible and robust networks that are implicated in almost every aspect of cellular physiology, ranging from protein degradation to receptor trafficking, DNA repair, cell-cycle progression, gene transcription, autophagy, and apoptosis.

Ubiquitin-like modifiers (ULMs)

In addition to ubiquitin, there is a small family of ubiquitin-like modifiers. The SUMOs and ISG15 are the other two ULMs that have been implicated as modifiers involved in HPV viral processes. There are four human SUMO genes encoding SUMOs 1-4. Small ubiquitinrelated modifier (Sumo-1) shares only 18% sequence identity with ubiquitin, whereas Nedd8 (neural precursor cell expressed developmentally downregulated-8) shares an 80% sequence homology with ubiquitin. It is not the sequence itself which makes these proteins function in a similar manner to ubiquitin, rather their apparent 3-D topologies. Both Sumo and Nedd8 have nearly superimposable 3-D structures compared to ubiquitin. Both proteins covalently attach to larger proteins, via a GlyGly motif at their c-termini and bind to the ε-amino group of lysine residues on acceptor proteins. Nedd-8 and Sumo-2 result in single addition of a molecule to the target, whereas Sumo 2/3 can result in poly-chains being conjugated to the substrate. ULM proteins also have a similar complex complement of E1 and E2 proteins that mediate their conjugation to target proteins.


1. Simon Thompson, Liam Loftus, Michelle Ashley et al. Ubiquitin-Proteasome System as a Modulator of Cell Fate. Curr Opin Pharmacol. 2008, 8(1): 90–95.

2. Van G. Wilson. The Role of Ubiquitin and Ubiquitin-Like Modification Systems in Papillomavirus Biology. Viruses.2014, 6: 3584-3611

3. Robert Layfield, R. John Mayer. The ubiquitin proteasome system in human disease. Biophysica Acta. 2008, 1782: 681–68

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