Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The . gov means it’s official. Federal government websites often end in . gov or . mil VSports app下载. Before sharing sensitive information, make sure you’re on a federal government site. .

Https

The site is secure V体育官网. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely. .

. 2006 Apr 11;103(15):5729-34.
doi: 10.1073/pnas.0504472103. Epub 2006 Mar 29.

Structure of human Wilson protein domains 5 and 6 and their interplay with domain 4 and the copper chaperone HAH1 in copper uptake

David Achila  1 , Lucia Banci, Ivano Bertini, Jennifer Bunce, Simone Ciofi-Baffoni, David L Huffman
Affiliations

Structure of human Wilson protein domains 5 and 6 and their interplay with domain 4 and the copper chaperone HAH1 in copper uptake

VSports注册入口 - David Achila et al. Proc Natl Acad Sci U S A. .

Abstract

Human Wilson protein is a copper-transporting ATPase located in the secretory pathway possessing six N-terminal metal-binding domains. Here we focus on the function of the metal-binding domains closest to the vesicular portion of the copper pump, i. e. , domain 4 (WLN4), and a construct of domains 5 and 6 (WLN5-6). For comparison purposes, some experiments were also performed with domain 2 (WLN2) VSports手机版. The solution structure of apoWLN5-6 consists of two ferredoxin folds connected by a short linker, and (15)N relaxation rate measurements show that it behaves as a unit in solution. An NMR titration of apoWLN5-6 with the metallochaperone Cu(I)HAH1 reveals no complex formation and no copper exchange between the two proteins, whereas titration of Cu(I)HAH1 with WLN4 shows the formation of an adduct that is in fast exchange on the NMR time scale with the isolated protein species as confirmed by (15)N relaxation data. A similar interaction is also observed between Cu(I)HAH1 and WLN2; however, the relative amount of the adduct in the protein mixture is lower. An NMR titration of apoWLN5-6 with Cu(I)WLN4 shows copper transfer, first to WLN6 then to WLN5, without the formation of an adduct. Therefore, we suggest that WLN4 and WLN2 are two acceptors of Cu(I) from HAH1, which then somehow route copper to WLN5-6, before the ATP-driven transport of copper across the vesicular membrane. .

PubMed Disclaimer

VSports app下载 - Conflict of interest statement

Conflict of interest statement: No conflicts declared.

Figures

Fig. 1.
Fig. 1.
Solution structure of apoWLN5–6. (A) Backbone superimposition of the 20 lowest-energy conformers of apoWLN5–6. The secondary structure elements are indicated: β-strands are cyan and α-helices are red. (B) Ribbon diagram of a representative structure of apoWLN5–6 in the same orientation as shown in A. Copper-binding residues (Cys-15, Cys-18, Cys-91, and Cys-94) are yellow. Residues belonging to the linker Met-72–Gly-80, which display NOEs with residues of the two domains, are magenta. The conserved negatively and positively charged residues, likely determining a preferential orientation between the two domains, also are labeled.
Fig. 2.
Fig. 2.
Cu(I) interaction studies of apoWLN5–6. (A) The NH crosspeaks, showing chemical shift δavg(HN) differences above the threshold of 0.03 ppm are shown in red on the backbone structure of apoWLN5–6. (B) The weighted average chemical shift differences δavg(HN) (i.e., {[(δH)2 + (δN/5)2]/2}1/2, where δH and δN are chemical shift differences for 1H and 15N, respectively) are shown. Chemical shifts differences are not reported for residues 14–17 and 90–93, because their 1H–15N cross-peaks are not observed for apo and/or Cu(I) forms. The secondary structure elements of apoWLN5–6 are reported at the top of the graph. The threshold is indicated with a horizontal red line.
Fig. 3.
Fig. 3.
Transfer of Cu(I) from Cu(I)WLN4 to apoWLN5–6. Superposition of the 1H–15N HSQC spectra of apoWLN5–6 (black) and apoWLN5–6 in the presence of unlabeled Cu(I)WLN4 at a 1: 2.5 molar ratio (red), showing the simultaneous presence of the signals of Val-19 and Val-95 in their Cu(I)- and apo-loaded state.
Fig. 4.
Fig. 4.
Complex formation between Cu(I)HAH1 and WLN4. (Right) The weighted average chemical shift differences δavg(HN) between 15N-Cu(I)HAH1 and the 1:1 15N-Cu(I)HAH1/apoWLN4 mixture. The secondary structure elements of Cu(I)HAH1 are reported at the top of the graph. The threshold is indicated with a horizontal red line. (Left) The NH crosspeaks showing chemical shift δavg(HN) differences above the threshold of 0.03 ppm are shown in red on the backbone structure of Cu(I)HAH1, whereas NH crosspeaks that disappear or significantly broaden are shown in blue. Cys-12 and Cys-15 are shown in yellow.
Fig. 5.
Fig. 5.
One of the paths for copper transfer from HAH1 to the N terminus of WLNP. (Step 1) First a complex forms between HAH1 and WLN4 (denoted 4), allowing rapid copper transfer between donor and acceptor. (Step 2) apoHAH1 diffuses away from WLN4, and, then (Step 3), WLN4 transfers copper to WLN5–6. Interdomain residue spacing is noted with numbers. Theoretically, the 57-aa linking region between WLN4 and WLN5–6 could allow the copper-binding site of WLN4 to access both domains of WLN5–6. Other targets of HAH1, i.e., WLN2, may function similarly.
See this image and copyright information in PMC

References

    1. Tanzi R. E., Petrukhin K., Chernov I., Pellequer J. L., Wasco W., Ross B., Romano D. M., Parano E., Pavone L., Brzustowicz L. M., et al. Nat. Genet. 1993;5:344–350. - PubMed
    1. Bull P. C., Thomas G. R., Rommens J. M., Forbes J. R., Cox D. W. Nat. Genet. 1993;5:327–337. - PubMed (VSports最新版本)
    1. Lutsenko S., Petris M. J. J. Membr. Biol. 2003;191:1–12. - PubMed
    1. Thomas G. R., Forbes J. R., Roberts E. A., Walshe J. M., Cox D. W. Nat. Genet. 1995;9:210–217. - VSports手机版 - PubMed
    1. Mercer J. F. Trends Mol. Med. 2001;7:64–69. - PubMed

Publication types

MeSH terms

LinkOut - more resources