Structural Diversity of Ubiquitin E3 Ligase

Abstract

The post-translational modification of proteins regulates many biological processes. Their dysfunction relates to diseases. Ubiquitination is one of the post-translational modifications that target lysine residue and regulate many cellular processes. Three enzymes are required for achieving the ubiquitination reaction: ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2), and ubiquitin ligase (E3). E3s play a pivotal role in selecting substrates. Many structural studies have been conducted to reveal the molecular mechanism of the ubiquitination reaction. Recently, the structure of PCAF_N, a newly categorized E3 ligase, was reported. We present a review of the recent progress toward the structural understanding of E3 ligases.

Keywords: X-ray crystallography; post-translational modification; structural biology; ubiquitin E3 ligase.

Figure 1

Schematic diagram of the ubiquitin conjugation system and the crystal structures of ubiquitin, E1, E2, and E3. (A) Schematic diagram of ubiquitin conjugation system. All molecules (ubiquitin, E1, E2, and E3) relating to the system are drawn in a circle and colored in orange, blue, gray, and purple, respectively. (B) The crystal structures of ubiquitin, E1, and E2 are drawn in a ribbon diagram and colored in orange, blue, and gray, respectively. PDB IDs are indicated under each structure.

Figure 2

Recognition of E2 by RING E3. (A) Schematic diagram of E2~Ub activation mechanism by RING E3. The structure of E2~ubiquitin prefers open conformations in which a ubiquitin molecule moves dynamically. RING E3 promotes a population shift toward closed conformations to stimulate the transfer activity of E2. (B) The crystal structures of the RING E3-UbcH5 complex. Ubiquitin, E2, and RING E3 are shown in a ribbon diagram and colored in orange, gray, and purple, respectively. PDB ID is shown under each structure. The position of catalytic cysteine is indicated as a pink circle. The Ile36 located on the ubiquitin surface contacting α2 of E2 is indicated as an orange circle.

Figure 3

Structures of classical and atypical E3 ligases. (A) The crystal structures of the RING E3, HECT E3, and RBR E3 domain are drawn in a ribbon diagram. The molecular name and PDB ID are shown under each structure. In the RING E3 structure, the RING domain is colored in purple, and the remaining structure is colored in pink. In HECT E3, N-lobe and C-lobe are colored in pink and purple, respectively. In RBR E3, RING1, IBR, and RING2 are colored in pink, pale purple, and purple, respectively. The linker region between IBR and RING1 is colored in gray. A pink circle indicates the position of catalytic cysteine. The schematic diagram of the ubiquitination mechanism of each E3 is drawn. (B) The crystal structures of atypical E3 ligase. The molecular name and PDB ID are shown under each structure. The structure of Ubl, E2, and E3 molecules are drawn in a ribbon diagram and colored in orange, gray, and purple, respectively.

Figure 4

Structure of PCAF_N domain. (A) Domain architectures of GCN5 and PCAF. The PCAF_N domain, the acetyltransferase domain (AT), and the bromo domain (Bromo) are indicated as a box and colored in purple, red, and blue, respectively. The amino acid sequence identities (%) are indicated on the right in which the number in parentheses indicates the amino acid region using sequence alignment. (B) The crystal structure of PCAF_N is drawn in a ribbon diagram in which the Zn region, the connective region, and the MSL-like region are colored in purple, pink, and pale purple, respectively. The PDB ID is indicated under the ribbon diagram. (C) The topology diagrams of the Zn-coordinating manner of the Ring region of the PCAF_N domain and Ring domain. The Zn-coordinating residues are indicated as the purple box. The PDB ID of the RING domain is shown. (D) The result of HMM logo in Pfarm (edited) is shown. The region around the residues coordinating Zn ions is shown. The seven ligand residues are indicated by an asterisk (*). The font size in HMM logo analysis shows the conservation of amino acids within the multiple sequence alignment. The amino acid drawn in a bigger size indicates that the amino acid is highly conserved among the protein family. GCN5 and PCAF have one amino acid insertion between the first and second cysteine residues. The underline indicates the corresponding sequence. (E) The structure around the Zn binding site of PCAF_N. Two Zn ions are drawn in the sphere model, and the coordination manner is indicated by the dashed black line. The Zn coordinating residues and highly conserved Trp residue (W118 in PCAF_N domain of mGCN5) are drawn in the stick model. W118 forms hydrogen bonding with the main chain carbonyl oxygens of E140 and G102. The two hydrogen bonds are indicated by a dashed purple line.

Figure 5

The domain architectures of PACF_N family proteins. The structured domains are shown. The names (species and proteins) are indicated on each domain architecture. The value in the first parenthesis is the number of amino acid residues of the presented protein. The value in the second parenthesis is the number of protein sequences harboring the same domain architecture.