title: Structural analysis of the UBA domain of X-linked inhibitor of apoptosis protein reveals different surfaces for ubiquitin-binding and self-association.
abstract: BACKGROUND
  Inhibitor of apoptosis proteins (IAPs) belong to a pivotal antiapoptotic protein family that plays a crucial role in tumorigenesis, cancer progression, chemoresistance and poor patient-survival. X-linked inhibitor of apoptosis protein (XIAP) is a prominent member of IAPs attracting intense research because it has been demonstrated to be a physiological inhibitor of caspases and apoptosis. Recently, an evolutionarily conserved ubiquitin-associated (UBA) domain was identified in XIAP and a number of RING domain-bearing IAPs. This has placed the IAPs in the group of ubiquitin binding proteins. Here, we explore the three-dimensional structure of the XIAP UBA domain (XIAP-UBA) and how it interacts with mono-ubiquitin and diubiquitin conjugates.
  
  
  RESULTS
  The solution structure of the XIAP-UBA domain was determined by NMR spectroscopy. XIAP-UBA adopts a typical UBA domain fold of three tightly packed α-helices but with an additional N-terminal 3(10) helix. The XIAP-UBA binds mono-ubiquitin as well as Lys48-linked and linear-linked diubiquitins at low-micromolar affinities. NMR analysis of the XIAP-UBA-ubiquitin interaction reveals that it involves the classical hydrophobic patches surrounding Ile44 of ubiquitin and the conserved MGF/LV motif surfaces on XIAP-UBA. Furthermore, dimerization of XIAP-UBA was observed. Mapping of the self-association surface of XIAP-UBA reveals that the dimerization interface is formed by residues in the N-terminal 3(10) helix, helix α1 and helix α2, separate from the ubiquitin-binding surface.
  
  
  CONCLUSIONS
  Our results provide the first structural information of XIAP-UBA and map its interaction with mono-ubiquitin, Lys48-linked and linear-linked diubiquitins. The notion that XIAP-UBA uses different surfaces for ubiquitin-binding and self-association provides a plausible model to explain the reported selectivity of XIAP in binding polyubiquitin chains with different linkages.
authors.affiliation: Department of Microbiology and State Key Laboratory for Emerging Infectious Diseases, The University of Hong Kong, Hong Kong SAR, People's Republic of China.
authors.forename: Man Kit
authors.initials: MK
authors.lastname: Tse
authors.forename: Sin Kam
authors.initials: SK
authors.lastname: Hui
authors.forename: Yinhua
authors.initials: Y
authors.lastname: Yang
authors.forename: Si-Tao
authors.initials: ST
authors.lastname: Yin
authors.forename: Hong-Yu
authors.initials: HY
authors.lastname: Hu
authors.forename: Bing
authors.initials: B
authors.lastname: Zou
authors.forename: Benjamin Chun Yu
authors.initials: BC
authors.lastname: Wong
authors.forename: Kong Hung
authors.initials: KH
authors.lastname: Sze
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D006801:Humans; D008239:Lysine / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D025821:Polyubiquitin / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D012996:Solutions; D013329:Structure-Activity Relationship; D013499:Surface Properties; D025801:Ubiquitin / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D012996:Solutions; D025801:Ubiquitin; D014452:Ubiquitins; D051636:X-Linked Inhibitor of Apoptosis Protein; C075986:diubiquitin conjugate; D025821:Polyubiquitin; D008239:Lysine