Crystal structure and activation mechanism of DR3 death domain

Abstract

Death receptor 3 (DR3) (a.k.a. tumor necrosis factor receptor superfamily 25) plays a key role in the immune system by activating nuclear factor kappa‐light‐chain‐enhancer of activated B cells signaling pathway. Here we present the crystal structures of human and mouse DR3 intracellular death domain (DD) at 2.7 and 2.5 Å resolutions, respectively. The mouse DR3 DD adopts a classical six‐helix bundle structure while human DR3 DD displays an extended fold. Though there is one‐amino‐acid difference in the linker between maltose‐binding protein (MBP) tag and DR3 DD, according to our self‐interaction analysis, the hydrophobic interface discovered in MBP–hDR3 DD crystal structure is responsible for both hDR3 DD and mDR3 DD homotypic interaction. Furthermore, our biochemical analysis indicates that the sequence variation between human and mouse DR3 DD does not affect its structure and function. Small‐angle X‐ray scattering analysis shows the averaged solution structures of both human and mouse MBP‐DR3 DD are the combination of different conformations with different proportion. Through switching to the open conformation, DR3 DD could improve the interaction with downstream element TNFR‐associated death domain (TRADD). Here we propose an activation‐dependent structural rearrangement model: the DD region is folded as the six‐helix bundles in the resting state, while upon extracellular ligand engagement, it switches to the open conformation, which facilitates its self‐association and the recruitment of TRADD. Our results provide detailed insights into the architecture of DR3 DD and the molecular mechanism of activation. Databases All refined structure coordinates as well as the corresponding structure factors have been deposited in the PDB under the accession codes http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5YGS, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5YEV, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5YGP, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5ZNY, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5ZNZ.

Biochemical characterization of SARS-CoV-2 nucleocapsid protein

“…Disuccinimidyl suberate (DSS, Pierce) was used to cross-link closely spaced surface-exposed active amino groups of interacting proteins. The experimental protocol is according to previous report [31].…”

Crystal structure of caspase-11 CARD provides insights into caspase-11 activation

“…The crystal structure of the caspase-11 CARD (residues 11–101) was determined at a resolution of 2.8 Å using X-ray crystallography with an N-terminal MBP as a crystallization tag (Table 1 ). This technique has been used successfully in the structure determination of several death domains 17 , 18 . As shown in Fig.…”

“…Death fold is generally packed loosely 19 and several death domain members have non-classic structures during activation. For example, DR3 DD H3 and H4 undergo a conformation switch when interacting with tumor necrosis factor receptor-associated death domain 17 ; FAD binding can induce the shift of helix H6 to H5 in Fas-associated protein with death domain, which forms a stem helix for the adaptor binding 22 . Therefore, the conformational changes of death domains are often associated with biological functions.…”

The Diversity and Similarity of Transmembrane Trimerization of TNF Receptors

“…Indeed, soluble TNFR1 CRD1 has been used to compete with CRD1-mediated receptor association, which inhibits receptor clustering and activation, as a new anti-arthritis treatment strategy ( Deng, 2007 ). In addition to the ECD, the self-interaction of the intracellular domains could also contribute to receptor clustering and this type of interaction has been well characterized, for example, for death receptors such as DR3 and Fas ( Scott et al, 2009 ; Wang et al, 2010 ; Yin et al, 2019 ).…”