Enterococcus hirae

Solioz, M. and Odermatt, A., Copper and silver transport by CopB-ATPase in membrane vesicles of Enterococcus hirae, J Biol Chem, 270 (16), 9217-9221, 1995. 

Figure 12.2 Copper chaperone function, (a) Copper homeostasis in Enterococcus hirae is affected by the proteins encoded by the cop operon. CopA, Cu1+-import ATPase CopB, Cu1+-export ATPase CopY, Cu1+-responsive repressor copZ, chaperone for Cu1+ delivery to CopY. (b) The CTR family of proteins transports copper into yeast cells. Atxlp delivers copper to the CPx-type ATPases located in the post Golgi apparatus for the maturation of Fet3p. (c) Coxl7p delivers copper to the mitochondrial intermembrane space for incorporation into cytochrome c oxidase (CCO). (d) hCTR, a human homologue of CTR, mediates copper-ion uptake into human cells. CCS delivers copper to cytoplasmic Cu/Zn superoxide dismutase (SOD1). Abbreviations IMM, inner mitochondrial membrane OMM, outer mitochondrial membrane PM, plasma membrane PGV, post Golgi vessel. Reprinted from Harrison et al., 2000. Copyright (2000), with permission from Elsevier Science.

Because of their importance in many enzymes, bacteria have had to develop uptake systems for copper and zinc. Copper uptake (and homeostasis, which is discussed in Chapter 8) has been extensively studied in the Gram-positive bacterium Enterococcus hirae. Two of the genes in the cop operon, copA and copB, encode membrane ATPases. An extracellular reductase CorA reduces Cu2+ to Cu+, which is taken up by CopA when copper is limiting. 

Solioz, M. and Stoyanov, J.V. (2003) Copper homeostasis in Enterococcus hirae, FEMS Microbiol. Rev., 27, 183-195. 

Magnani, D. and Solioz, M. (2005) Copper chaperone cycling and degradation in the regulation of the cop operon of Enterococcus hirae, BioMetals, 18, 407-412. 

Murata, T., Arechaga, I., Fearnley, I. M., Kakinuma, Y., Yamato, I., and Walker, J. E. (2003). The membrane domain of the Na+-motive V-ATPase from Enterococcus hirae contains a heptameric rotor. J. Biol. Chem. 278, 21162-21167. 

Takase, K., Kakinuma, S., Yamato, I., Konishi, K., Igarashi, K., andKakinuma, Y. (1994). Sequencing and characterization of the ntp gene cluster for vacuolar-type Na(+)-translocating ATPase of Enterococcus hirae. J. Biol. Chem. 269, 11037-11044. 

Copper ion homeostasis in prokaryotes involves Cu ion efflux and sequestration. The proteins involved in these processes are regulated in their biosynthesis by the cellular Cu ion status. The best studied bacterial Cu metalloregulation system is found in the gram-positive bacterium Enterococcus hirae. Cellular Cu levels in this bacterium control the expression of two P-type ATPases critical for Cu homeostasis (Odermatt and Solioz, 1995). The CopA ATPase functions in Cu ion uptake, whereas the CopB ATPase is a Cu(I) efflux pump (Solioz and Odermatt, 1995). The biosynthesis of both ATPases is regulated by a Cu-responsive transcription factor, CopY (Harrison et al., 2000). In low ambient Cu levels Cop Y represses transcription of the two ATPase genes. On exposure to Cu(I), CopY dissociates from promoter/operator sites on DNA with a for Cu of 20 jlM (Strausak and Solioz, 1997). Transcription of copA and copB proceeds after dissociation of CuCopY. The only other metal ions that induce CopY dissociation from DNA in vitro are Ag(I) and Cd(II), although the in vivo activation of copA and copB is specihc to Cu salts. The CuCopY complex is dimeric with two Cu(I) ions binding per monomer (C. T. Dameron, personal communication). The structural basis for the Cu-induced dissociation of CopY is unknown. Curiously, CopY is also activated in Cu-dehcient cells, but the mechanism is distinct from the described Cu-induced dissociation from DNA (Wunderh-Ye and Solioz, 1999). 

Fig. 1. Comparison of structures of the apo-CopZ and Hg-Atxl metallochaperones irom. Enterococcus hirae and Saccharoniyces cerevisiae, respectively (Rosenzweigeta/., 1999 Wimmer et al., 1999). In the HgAtxl structure the Hg(ll) atom (shown as a dark ball) is ligated by two cysteines (the sulfurs in the side chains are shown as smaller balls). The coordination of the Hg(II) is linear a similar coordination geometry is expected for Cu(I). In the CopZ structure the two corresponding cysteinyl residues shown by arrows are not in the proper orientation to ligate Cu(I). A limited structural rearrangement is expected in the loop to permit linear coordination as seen in Hg-Atxl.

Fig. 3. Comparison of the membrane topology of a CPx-type ATPase and a nonheavy metal ATPase. Shown are CopB of Enterococcus hirae and the Ca -ATPase of sarcoplasmic reticnlnm. Helices common to both types of ATPases are in gray and helices nniqne to one type of ATPase are in black. Key seqnence motifs are indicated in single-letter amino acid code. In the center of the hgnre, the approximate locations of the three cytoplasmic domains. A, P, and N, are indicated. MBD, metal-binding domain containing repeat metal-binding sites TGE, conserved site in transdnction domain (A) CPx, pntative copper-binding site DKTGT, phosphorylation site in domain P HP, motif of nnknown function, probably in domain N GDG, nucleotide-binding site residues in domain N.

Fig. 5. Copper homeostasis in Enterococcus hirae. Under copper-limiting conditions, copper is pumped into the cell by CopA. The CopZ copper chaperone picks up copper at this site of entry. Under physiological copper conditions, Zn(II)CopY binds to the promoter and represses transcription of the cop operon. Under conditions of copper excess, Cu-CopZ donates Cu(I) to CopY, which leads to the replacement of the Zn(II), loss of DNA-binding affinity, and ultimately synthesis of the operon products. Excess copper is secreted by the CopB efflux pump. The substrate for this pump may be a copper-glutathione (GSH) complex, rather than Cu-CopZ.

Fig. 6. Occurrence of the CXCX(4 5) CXC consensus motif. CopY, cop operon repressor protein from Enterococcus hirae Mad, transcription factor for the Ctrl copper transporter of Saccharomyces cerevisiae AMTl, transcription factor for metal-lothionein from Candida albicans ACEl, transcription factor for metallothionein from Sa. cerevisiae Grisea, MACl orthologue of Podospora anserina MT-2 p-domain, N-terminal domain of human metallothionein-2.

Enterococcus hirae CopA and CopZ Odermatt and Solioz, 1995... 

Fig. 5. Three-dimensional structures of the (iaPPaP proteins oftheAtxl-like family. MXCXXC motif residues are hoxed. The Protein Data Bank (pdh) code for each structure is in parentheses, (a) NMR structure of Shigella flexneri Hg(II)MerP (Steele and Opella, 1997). (h) X-ray structure oiSaccharamyces cerevisiae Hg(II)Atxl. K24, K28, K59, and K62, side chains important in the recognition of the Ccc2a target domain, are shown outside of the hox (see text) (Rosenzweig et al., 1999). (c) X-ray structure of human Cu(I)Hahl. R21, K25, K56, and K57, side chains important in the recognition of the fourth N-terminal domain of the Menkes protein, are shown outside of the box (Wernimont et al., 2000). (d) NMR structure oi Enterococcus hirae apoCopZ (Wimmer et al., 1999). (e) NMR structure of human Ag(l)Mnk4, the fourth domain of the...

In the gram-positive bacteria Enterococcus hirae, copper homeostasis is maintained by the cop operon, consisting of the copA, copB, copY, and copZ... 

Odermatt et al., 1993) proteins of Pseudomonas syringae and Enterococcus hirae, respectively. An example of this sequence, arbitrarily chosen from Xanthomonas campestris, is... 

Odermatt, A, Suter, H., Krapt, R., and Solioz, M. (1993). Primary structure of two P-type ATPases involved in copper homeostasis in Enterococcus hirae. J. Biol. Chem. 268, 12775-12779. 

Lactobacillus brevis Enterococcus faecalis Enterococcus hirae Lactococcus lactis Enterococcus faecium... 

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