Ubiquitin ligase, also known as E3 ubiquitin ligase, is an enzyme that binds a ubiquitin molecule to a certain lysine of a protein of interest. In general, ubiquitin ligase can polyubiquitinate a protein of interest, that is, add multiple ubiquitin molecules to form a polyubiquitin chain; while a protein with a polyubiquitin chain can be recognized by the proteasome. However, in some cases, ubiquitin ligase only binds a ubiquitin molecule to the protein of interest; this monoubiquitinated protein is not degraded by the proteasome, but its localization or function in the cell may change. For example, a monoubiquitinated protein can change its position by binding to other proteins with a ubiquitin-binding domain.
Figure 1. Crystal structure of Ubiquitin ligase.
Ubiquitin ligase is the third enzyme in the protein ubiquitination pathway (E3). The first two enzymes are ubiquitin activating enzyme (E1) and ubiquitin cross-linking enzyme (E2). The ubiquitin ligase E3 is divided into the HECT (homologous to E6APC terminus) family and the RING-finger family. The former also has the HECT domain, in which the conserved Cys residue can form a thioester bond with the ubiquitin carried by E2, E2 will first Ubiquitin is passed to E3, which is then presented to the substrate by E3. The E3 of the RING-finger family contains a similar E2 binding domain, which acts as a bridge to transfer activated ubiquitin from E2 directly to the target protein, which itself does not interact with ubiquitin.
Ubiquitin activating enzyme (E1)
Ubiquitin activating enzymes are involved in the first step in catalyzing the ubiquitination reaction, which, among other things, can target proteins for degradation by the proteasome. This covalent attachment of ubiquitin or a ubiquitin-like protein to a target protein is the primary mechanism for regulating protein function in eukaryotes. Many processes, such as cell division, immune response, and embryonic development, are also regulated by post-translational modifications of ubiquitin and ubiquitin-like proteins. At the beginning of the ubiquitination cascade, the E1 enzyme binds to ATP-Mg2+ and ubiquitin, catalyzing the C-terminal acylation of ubiquitin. Next, the catalytic cysteine on the E1 enzyme attacks the ubiquitin-AMP complex by acyl substitution, while forming a thioester bond and an AMP leaving group. Finally, the E1 to ubiquitin complex transfers ubiquitin to the E2 enzyme by transthioesterification, in which the E2 catalyzed cysteine attacks the back side of the E1 to ubiquitin complex. However, since both the E1 and E2 enzymes form an intermediate complex, both enzymes undergo a series of conformational changes to bind to each other, and therefore, the sulfur transfer esterification process is very complicated. Throughout the mechanism, the E1 enzyme binds to two ubiquitin molecules. Although this ubiquitin is similarly adenylated, it does not form the same thioester complex as previously described. The function of secondary ubiquitin is still largely unknown, but it is believed that it may promote conformational changes seen in the E1 enzyme during transthioesterification.
Figure 2. Crystal structure of human ubiquitin activating enzyme E1 (Uba1) in complex with ubiquitin.
Ubiquitin ligase family
1. Late phase promoting complex
The late-promoting complex/cell cycle (APC/C) is a large E3 ligase that mediates ubiquitin-dependent proteolysis of cell cycle regulatory proteins to control events during replication and cell division. In the process of ubiquitination and degradation of M-phase cyclin, a complex with E3 activity exists, which becomes a late-promoting complex, which can combine ubiquitin and substrate and degrade by proteasome. APC is ubiquitin ligase, E3) complex. E3 is usually a complex composed of multiple subunits. APC stimulates the E2-ubiquitin complex to bind to the mitotic cyclin disruption framework, and then stimulates ubiquitin with the lysine at the C-terminus of the disruption cassette. The residues are combined and this process continues to circulate to ubiquitin.
2. SCF complex
The complex (or SCF complex) comprising Skp, Cullin, F-box is a polyprotein E3 ubiquitin ligase complex that catalyzes the ubiquitination of proteins intended for proteasomal degradation. It plays an important role in the ubiquitination of proteins involved in the cell cycle, and also marks a variety of other cell-damaging proteins.
1. Schulman BA.; et al. Ubiquitin-like protein activation by E1 enzymes: the apex for downstream signalling pathways. Nature Reviews Molecular Cell Biology. 2009, 10 (5): 319–31.