title: Characterisation of assembly and ubiquitylation by the RBCC motif of Trim5α.
abstract: The post-entry restriction factor Trim5α blocks infection of retroviral pathogens shortly after the virus gains entry to the cell, preventing reverse transcription and integration into the host genome. Central to the mechanism of restriction is recognition of the lattice of capsid protein that forms the inner-shell of the retrovirus. To recognise this lattice, Trim5α has been shown to assemble into a large hexagonal array, complementary to the capsid lattice. Structures of the Trim5α coiled-coil region reveal an elongated anti-parallel dimer consistent with the edges of this array placing the Bbox domain at each end of the coiled-coil to facilitate assembly. To investigate the nature of this assembly we have designed and characterised a monomeric version of the TRIM RBCC motif with a truncated coiled-coil. Biophysical characterisation by SEC-MALLS, AUC, and SAXS demonstrate that this construct forms compact folded domain that assembles into a trimer that would support the formation of a hexagonal lattice. Furthermore, the RING domain and elements of the coiled-coil region are shown to contribute to assembly. Ubiquitylation assays demonstrate that this assembly increases ubiquitylation activity providing a link from recognition of the capsid lattice and assembly to the activation of innate immune signalling and restriction.
authors.affiliation: School of Biological Sciences, University of Auckland, Auckland, New Zealand.
authors.forename: Jeremy R
authors.initials: JR
authors.lastname: Keown
authors.affiliation: School of Biological Sciences, University of Auckland, Auckland, New Zealand.
authors.forename: Joy X
authors.initials: JX
authors.lastname: Yang
authors.affiliation: School of Biological Sciences, University of Auckland, Auckland, New Zealand.
authors.forename: Jordan
authors.initials: J
authors.lastname: Douglas
authors.affiliation: School of Biological Sciences, University of Auckland, Auckland, New Zealand.|Associate Investigator Maurice Wilkins Centre for Molecular Biodiscovery, New Zealand.
authors.forename: David C
authors.initials: DC
authors.lastname: Goldstone
mesh_terms: D000090103:Antiviral Restriction Factors; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D054730:Protein Interaction Domains and Motifs; D000070584:Tripartite Motif Proteins; D044767:Ubiquitin-Protein Ligases; D054875:Ubiquitination
chemical_list: D000090103:Antiviral Restriction Factors; D002352:Carrier Proteins; D000070584:Tripartite Motif Proteins; C477346:TRIM5 protein, human; D044767:Ubiquitin-Protein Ligases