title: Copper-binding properties of the BIR2 and BIR3 domains of the X-linked inhibitor of apoptosis protein.
abstract: The X-linked inhibitor of apoptosis protein (XIAP) is a zinc metalloprotein that has recently been implicated in copper homeostasis. XIAP mediates apoptosis via the inhibition of caspase enzymes through multiple baculovirus IAP repeat (BIR) domains, wherein zinc is coordinated by three cysteine amino acids and one histidine amino acid. XIAP binds copper ions directly at one or more unspecified sites, indicating that the protein may function as a copper sensor. We report the copper-binding properties of an XIAP construct containing the BIR2 and BIR3 domains. Absorption and emission spectroscopic measurements show that XIAP exhibits only a low-to-moderate affinity for Cu(II), but a strong affinity for Cu(I). Cu(I) is observed to bind at multiple sites within the BIR2 and BIR3 domains, including the CXXC motifs of the zinc structural sites and multiple BIR2 surface sites. Mutagenesis-based experiments reveal that surface cysteine residues mediate binding in the BIR2 domain and induce protein oligomerization under elevated copper concentrations. These results constitute the first spectroscopic evidence of copper-XIAP interactions.
authors.affiliation: Department of Chemistry, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105, United States.
authors.forename: Yiwen
authors.initials: Y
authors.lastname: Liang
authors.affiliation: Department of Chemistry, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105, United States.
authors.forename: Patrick M
authors.initials: PM
authors.lastname: Ewing
authors.affiliation: Department of Chemistry, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105, United States.
authors.forename: Willem J
authors.initials: WJ
authors.lastname: Laursen
authors.affiliation: Department of Chemistry, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105, United States.
authors.forename: Valerie T
authors.initials: VT
authors.lastname: Tripp
authors.affiliation: Department of Chemistry, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105, United States.
authors.forename: Shubha
authors.initials: S
authors.lastname: Singh
authors.affiliation: Department of Chemistry, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105, United States. Electronic address: splank@macalester.edu.
authors.forename: Kathryn E
authors.initials: KE
authors.lastname: Splan
mesh_terms: D000595:Amino Acid Sequence; D003300:Copper / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000737:chemistry / Q000378:metabolism*
chemical_list: D051636:X-Linked Inhibitor of Apoptosis Protein; D003300:Copper
keywords: Copper homeostasis; Cu(I); Cysteine mutagenesis; Inhibitor of apoptosis protein; Metal binding