Disease relevance of ATOX1

• Homologs of the copper transporting ATPase ATP7B, defective in Wilson disease, and the copper chaperone ATOX1 were potential candidates, but both have been excluded [1].
• Characterization of the interaction between the Wilson and Menkes disease proteins and the cytoplasmic copper chaperone, HAH1p [2].
• The human metallochaperone HAH1 has been produced in Escherichia coli with four additional amino acids at the C-terminus and characterized in solution by NMR spectroscopy, both with and without copper(I) [3].
• The copper metallochaperone Atx1 of chloroplast-related cyanobacteria (ScAtx1) engages in bacterial two-hybrid interactions with N-terminal domains of copper-transporting ATPases CtaA (cell import) and PacS (thylakoid import) [4].

High impact information on ATOX1

• Structure of human Wilson protein domains 5 and 6 and their interplay with domain 4 and the copper chaperone HAH1 in copper uptake [5].
• The Hah1 metallochaperone protein is implicated in copper delivery to the Menkes and Wilson disease proteins [6].
• The 1.8 A resolution structure of CuHah1 reveals a copper ion coordinated by Cys residues from two adjacent Hah1 molecules [6].
• An extended hydrogen bonding network, unique to the complex of two Hah1 molecules, stabilizes the metal binding sites and suggests specific roles for several conserved residues [6].
• Interaction of the copper chaperone HAH1 with the Wilson disease protein is essential for copper homeostasis [7].

Chemical compound and disease context of ATOX1

• Studies with a glutathione S-transferase-HAH1 fusion protein demonstrated direct protein-protein interaction between HAH1 and the Wilson disease protein, which required the cysteine copper ligands in the amino terminus of HAH1 [7].
• Metallochaperone Atox1 transfers copper to the NH2-terminal domain of the Wilson's disease protein and regulates its catalytic activity [8].

Biological context of ATOX1

• The human phenotype resulting from mutations in ATOX1 remains unidentified [9].
• Kinetic analysis of the interaction of the copper chaperone Atox1 with the metal binding sites of the Menkes protein [10].
• Taken together, these data provide a mechanism for the function of HAH1 as a copper chaperone in mammalian cells and demonstrate that this protein is essential for copper homeostasis [7].
• The incubation with apo-Atox1 results in the removal of copper from the metalated N-WNDP and apparent down-regulation of WNDP activity [8].

Anatomical context of ATOX1

• In this current study, HAH1 was detected in lysates from multiple human cell lines and tissues as a single-chain protein distributed throughout the cytoplasm and nucleus [7].

Associations of ATOX1 with chemical compounds

• Alteration of either the Met or Thr residue of the HAH1p MTCXXC motif has no observable effect on the copper-dependent interaction, whereas alteration of either of the two Cys residues abolishes the interaction [2].
• Mutation of any one of the HAH1p C-terminal Lys residues (Lys(56), Lys(57), or Lys(60)) to Gly does not affect the interaction, although deletion of the 15 C-terminal residues abolishes the interaction [2].
• The in vitro HAH1/WDp interaction is metalospecific; HAH1 preincubated with Cu(2+) or Hg(+) but not with Zn(2+), Cd(2+), Co(2+), Ni(3+), Fe(3+), or Cr(3+) interacted with WDp [2].
• Although a substantial amount of structural data exist for HAH1 and its yeast and bacterial homologues, details of the copper coordination remain unclear and suggest the presence of two protein-derived cysteine ligands and a third exogenous thiol ligand [11].
• When GSH or dithiothreitol was added to the chaperone during the reconstitution procedure, the resulting Cu(I)- HAH1 remained two-coordinate, whereas the addition of the phosphine during reconstitution elicited a three-coordinate species [11].

Physical interactions of ATOX1

• Atox1 delivers copper to the secretory pathway and docks with either copper-transporting ATPase ATP7B in the liver or ATP7A in other cells [12].
• We have used isothermal titration calorimetry to measure the association constant (K(a)) and stoichiometry (n) values of Cu(I) binding to the WND metal-binding domains and to their metallochaperone Atox1 [13].
• HAH1 is a copper-binding protein with distinct amino acid residues mediating copper homeostasis and antioxidant defense [14].

Regulatory relationships of ATOX1

• Our data also suggest that Atox1 can regulate the copper occupancy of WNDP [8].
• The HAH1 gene is expressed in haematological tissues and plays a role in antioxidant defence [15].

Other interactions of ATOX1

• Atx1 and ATOX1 both contain an MXCXXC motif that is also present in Ccc2 (two motifs) and ATP7A/B (six motifs) [16].
• In addition, we speculate on how copper binding may regulate the activity and intracellular distribution of WNDP, and what role the human copper chaperone Atox1 may play in these processes [17].
• The HAH1 gene maps immediately adjacent to the SPARC gene at 5q32, and is flanked by the genetic markers D5S1838 and D5S1419 [15].
• Competition experiments using a copper chelator revealed that MBS2 retained copper much better than Atox1, and this may facilitate the metal transfer process [18].
• These results identify HAH1 as a novel ubiquitously expressed protein, which may play an essential role in antioxidant defense and copper homeostasis in humans [19].

Analytical, diagnostic and therapeutic context of ATOX1

• Although the yeast two-hybrid assay failed to show an interaction of Atox1 with MBS5/6, SPR analysis clearly demonstrated a copper-dependent binding with all six MBSs highlighting the power and sensitivity of SPR as compared with other, more indirect methods like the yeast two-hybrid system [10].
• Here we investigated interactions of Atox1 with wild-type and mutated pairs of the MBSs of MNK using two different methods: yeast two-hybrid analysis and real-time surface plasmon resonance (SPR) [10].
• The study was carried out through titrations involving HAH1 and either the second or the fifth soluble domains of ATP7A (MNK2 and MNK5, respectively), in the presence of copper(I) [20].
• Consistent with this model, in vitro studies with recombinant HAH1 directly demonstrated binding of Cu(I), and site-directed mutagenesis identified these cysteine residues as copper ligands [14].
• Fine physical mapping of the HAH1 gene within this genomic interval was then performed by screening YAC and BAC contigs spanning the critical region of the 5q- syndrome using PCR amplification [15].

References