Copper metallochaperones

Abstract

The current state of knowledge on how copper metallochaperones support the maturation of cuproproteins is reviewed. Copper is needed within mitochondria to supply the Cu(A) and intramembrane Cu(B) sites of cytochrome oxidase, within the trans-Golgi network to supply secreted cuproproteins and within the cytosol to supply superoxide dismutase 1 (Sod1). Subpopulations of copper-zinc superoxide dismutase also localize to mitochondria, the secretory system, the nucleus and, in plants, the chloroplast, which also requires copper for plastocyanin. Prokaryotic cuproproteins are found in the cell membrane and in the periplasm of gram-negative bacteria. Cu(I) and Cu(II) form tight complexes with organic molecules and drive redox chemistry, which unrestrained would be destructive. Copper metallochaperones assist copper in reaching vital destinations without inflicting damage or becoming trapped in adventitious binding sites. Copper ions are specifically released from copper metallochaperones upon contact with their cognate cuproproteins and metal transfer is thought to proceed by ligand substitution.

Figure 1

(a) Copper catalyzes production of hydroxyl radicals via the Fenton reaction. (b) Copper has a tendency to form stable complexes relative to other essential divalent metals, and hence it is at the top of the Irving-Williams series. (b) Monovalent copper is also competitive, forming tight protein complexes.

Figure 2

Copper is passed (green arrows) to cuproproteins (dark green circles), including metallothioneins Crs5 and Cup1, in for example Saccharomyces cerevisiae, by copper metallochaperones (pale green circles, or ovals if membrane associated). Copper permeases (white ovals) import copper into the cell or deliver Cu(I) to compartments, such as the trans-Golgi network where secreted cuproproteins (such as Fet3) acquire copper. Fet3 oxidizes ferrous ions (curved brown arrow) to provide substrate for high-affinity iron import by Ftr1. P₁-type ATPases (such as Ccc2) have one or more soluble N-terminal domains, which engage in regulatory interactions with copper metallochaperones (such as Atx1), but additional Cu(I) donation (hence two arrows) may supply the trans-membrane regions for Cu(I) transport. Metallation of the cytochrome c oxidase complex (CcO) in mitochondria (OM, outer membrane of mitochondria) involves a pathway via Cox11 for the Cu_B site of the Cox1 subunit, embedded in the inner membrane (IM), and via Sco1 for the Cu_A site of Cox2, protruding into the intermembrane space (IMS). Cox17 supplies both pathways. Nuclear encoded mitochondrial proteins, such as Cox17 and Ccs1, are imported across the OM unfolded (pale green line) via the TOM translocase then captured in the IMS, following introduction of disulfide bonds (SS) through the actions of Mia40. A sulfhydryl oxidase Erv1 generates a reactive disulfide on Mia40. A small copper ligand (CuL) supplies Cu(I) (dashed arrows) to the IMS copper metallochaperones.

Figure 3

(a) NMR solution structural model [Protein Data Bank (PDB) code 2GGP] of a heterodimer of the S. cerevisiae copper metallochaperone Atx1 and one of two N-terminal cytosolic domains of its cognate P₁-type copper-transporting ATPase Ccc2. Cysteinyl (yellow) thiol coordinated Cu(I) (sphere) stabilizes a transient complex between similar βαββαβ (ferredoxin)-structural folds (red, α-helix; blue, β-strand). (b) The interacting faces of Ccc2a and Atx1 have complementary (red, negative; blue, positive) electrostatic potentials.

Figure 4

Visualization of the Cys-Xaa-Xaa-Cys motifs taken from NMR solution structures (Protein Data Bank codes indicated) of the apo-, Cu(I)-, and heterodimeric forms of Atx1 and Ccc2a to represent the transfer of Cu(I) from Atx1 to Ccc2a via ligand-exchange reactions.

Figure 5

Copper metallochaperones for CcO. (a) Disulfide bonds between two pairs of cysteine residues (yellow) on antiparallel α-helices (red) create a helical hairpin of Cox17 [Protein Data Bank (PDB) code 2RNB]. Cu(I) is coordinated to a pair of cysteine residues. (b) Sco1 is tethered to the inner membrane (IM) (hydrophobic α-helix not shown) with a thioredoxin fold (β-strand, dark blue; histidine, pale blue) in the intermembrane space (PDB 2GQM). (c) Cox11 is tethered to the IM (hydrophobic α-helix not shown) with an immunoglobulin-like β-barrel in the IMS (PDB 1SPO).

Figure 6

Visualization of metal-binding motifs taken from structures (Protein Data Bank codes indicated) of the apo and Cu(I) forms of Cox17 and Sco1 to represent the transfer of Cu(I) from Cox17 to Sco1 and, potentially, to represent the coincident transfer of two electrons when apo-Sco1 is oxidized.