Third BIR domain of XIAP binds to both Cu(II) and Cu(I) in multiple sites and with diverse affinities characterized at atomic resolution

Abstract

The X-chromosome linked inhibitor of apoptosis, XIAP, is mainly known as the inhibitor of caspases by direct interaction with caspases with its baculoviral IAP repeat (BIR) domains. XIAP has three BIR domains and each BIR domain contains a zinc binding site, normally known as zinc finger motif. Recent studies showed that XIAP is involved in copper homeostasis in cells and the BIR domains bind copper ion. However, structural details of the second and third BIR domain, BIR2 and BIR3, in XIAP, with copper as well as the binding modes are not known. In the present work we characterize the structural properties of BIR3 in solution by high resolution NMR and other biophysical techniques. The interaction of BIR3 with copper both in vitro and in cell lysates was analyzed. Our results show that BIR3 is able to form stable complexes both with Cu(II) and Cu(I), whereas zinc binding site is not affected and zinc retains tightly bound in the zinc finger during these interactions. Surprisingly, BIR3 has multiple binding sites for Cu(II) and Cu(I) but with varied binding affinities. In addition, the solvent exposed Cys351 is readily oxidized by Cu(II) resulting an intermolecular disulfide bond either between two BIR3 molecules or a mixed disulfide bond with glutathione in cell lysates.

The expanded inhibitor of apoptosis gene family in oysters possesses novel domain architectures and may play diverse roles in apoptosis following immune challenge

“…To assess the potential functional diversity of oyster IAPs, oyster BIR sequences were compared to the IAP-defining BIR Type I and Type II BIR domains from the best studied reference model organisms to include a range of BIR domain diversity across taxa in vertebrates ( Homo sapiens , Mus musculus, Danio rerio ), and one invertebrate D. melanogaster [ 46 ]. In D. melanogaster and H. sapiens , BIR domains are characterized by 15 conserved amino acids forming a central 3-stranded antiparallel β-sheet (β1–3) surrounded by 5 α-helices (α1–5), with four critical residues stabilizing a central zinc atom: Histidine (H77) and three Cysteine residues (C57, C60, C84) [ 46 – 48 ] (Fig. 3 a).…”

“…A final, but potentially non-functional, novel BIR type in C. gigas and C. virginica was identified by hydrophilic Threonine amino acid substitution at the first coordinating Cysteine residue (C57) of this zinc-binding structural hot spot [ 46 – 48 ]. Though this substitution is not predicted to alter protein secondary structure, loss of this Cysteine may result in decreased ability for these domains to coordinate with Zinc [ 46 – 48 ]; therefore, this domain is referred to as Non-Zinc Binding (NZBIR) here (Fig. 3 b).…”

Review of cancer cell resistance mechanisms to apoptosis and actual targeted therapies

“…However, only XIAP can directly inhibit caspases, whereas cIAP only prevents the formation of proapoptotic signaling complexes in the extrinsic apoptotic pathway 111 . XIAP contains three baculoviral IAP repeats (BIR) and contains an associated ubiquitin domain 112 . The BIR domain serves as a protein‐protein interaction domain for the direct binding and inhibition of caspases‐3, ‐7, and ‐9 113 .…”

“…111 XIAP contains three baculoviral IAP repeats (BIR) and contains an associated ubiquitin domain. 112 The BIR domain serves as a protein-protein interaction domain for the direct binding and inhibition of caspases-3, -7, and -9. 113 The BIR3 domain inhibits caspase-9 and the binding region between the BIR1 and BIR2 domains is responsible for the inhibition of caspase-3 and -7.…”

Cuprous ions can disrupt the structure and functions of the RING finger domain of RNF11

“…Nevertheless, it has been reported that Cu(I), an isoelectronic ion of Zn(II), could substitute zinc coordination in several zinc finger proteins, consequently altering the structure and function of the proteins. [15][16][17][18] On the other hand, cellular analyses indicate that RING finger proteins, such as XIAP, regulate copper homeostasis, and this process is associated with cell survival. 19,20 Copper homeostasis is crucial for cell proliferation; upregulation of the copper level often occurs in tumorigenesis, 21 whereas excessive cellular copper is harmful to cells.…”