西亚试剂：Structural insights into the TRIM family of ubiquitin E3 li

TRIM proteins play important roles in a wide range of biological processes, including cell proliferation, differentiation, development, apoptosis, oncogenesis and innate immunity1,2. The N-terminal regions of all TRIM proteins contain a RING finger domain followed by one or two B-box domains and a coiled-coil domain (CCD). The RING-finger domain comprises conserved cysteine and histidine residues that bind two zinc atoms in a 'cross-brace' arrangement, and is essential for recruiting the ubiquitin-charged ubiquitin-conjugating enzymes (E2~Ub). The B-box domains, differing in both the number and spacing of the conserved cysteine and histidine residues3, are typically composed of B1 and B2 domains, but some TRIM members only contain a B2 domain. CCD following the B-box domains has been proposed to mediate protein-protein interactions, particularly homomeric and heteromeric interactions by forming intertwining helices among TRIM family and other proteins4,5.

Unlike some ubiquitin ligases that form multi-subunit complexes and catalyze ubiquitination with the help of other subunits6, TRIM proteins can exert their ubiquitin ligase activity as single-component ubiqutin ligases7,8. However, the underlying mechanism remains unknown. To understand how the highly conserved CCDs contribute to TRIM-catalyzed unbiquitination, we determined a crystal structure of the CCD (residues 143 to 321) of human TRIM69 at 2.15 Å resolution (Figure 1A, 1B and Supplementary information, Figure S1). The structure reveals that TRIM69 CCD forms a homodimer in an anti-parallel orientation (Figure 1B). Each monomer contains three α-helices (α1-α3). The N-terminal 112 residues (152 to 263) form the α1 helix with the length of about 168 Å. The α2 helix containing residues 266 to 288 folds back to form a 3-helix bundle with the two α1 helices of the dimer (Figure 1B and 1C). The short α3 helix (residues 305 to 318) is almost perpendicular to the long axis of the two long α1 helices (Figure 1C and Supplementary information, Figure S2A). In TRIM69, a conserved PRY/SPRY domain (also known as B30.2 domain) follows the CCD. Structural comparison of homologous PRY/SPRY domains in different species reveals that they share a highly similar fold (backbone rmsd of 0.65–1.25 Å), despite their low sequence identity (ranging from 15.7% to 24.7%) (Supplementary information, Figure S2B and S2C). Interestingly, most of them contain a short α-helix at their N-termini (Supplementary information, Figure S2B), which appears equivalent to the α3 helix in our CCD structure. Indeed, structural modeling shows that the configuration of α3 allows the PRY/SPRY domain to protrude from the rod-like coiled coil without steric clashing (Figure 1B, 1C and Supplementary information, Figure S2A).

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