CopH from Cupriavidus metallidurans CH34. A novel periplasmic copper-binding protein.
The copH gene is one of the 19 open reading frames (ORFs) found in the cop cluster borne by the large plasmid pMol30 in Cupriavidus metallidurans CH34. The entire cluster is involved in detoxification of copper from the cytoplasm as well as from the periplasm. The function of the corresponding protein, CopH, is not yet clear, but it seems to be involved in the late response phase. We have cloned copH and overproduced and purified the corresponding protein. CopH is rather unique as only one paralog can be found in the databases. It is a dimeric protein with a molecular mass of 13 200 Da per subunit and located in the periplasm. The metal binding properties of CopH were examined by using a series of techniques such as UV-visible spectroscopy, circular dichroism (CD), electron paramagnetic resonance (EPR), surface plasmon resonance (SPR), mass spectrometry, and nuclear magnetic resonance (NMR). All together, the corresponding data are consistent with a dimeric protein containing one metal-binding site per subunit. These sites have a high affinity for Cu(II) but can also bind zinc or nickel. CopH does not contain any cysteines or methionines but contains two histidines. EPR and UV-visible features are consistent with the presence of Cu(II) type 2 centers in a nitrogen ligand field. SPR data confirm the involvement of the histidine residues in copper binding. CD and NMR data reveal that CopH is partially unfolded.
Sendra
Véronique
V
Laboratoire des Protéines Membranaires, Institut de Biologie Structurale-Jean-Pierre Ebel, UMR 5075 CNRS-CEA-UJF, 41, rue Jules Horowitz, 38027 Grenoble Cedex, France.
Cannella
Dominique
D
Bersch
Beate
B
Fieschi
Franck
F
Ménage
Stéphane
S
Lascoux
David
D
Covès
Jacques
J
D000595:Amino Acid Sequence; D042521:Burkholderiaceae / Q000737:chemistry*; D002352:Carrier Proteins / Q000302:isolation & purification*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003300:Copper / Q000378:metabolism*; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D019897:Periplasm / Q000737:chemistry; D016415:Sequence Alignment; D013053:Spectrophotometry; D020349:Surface Plasmon Resonance
D002352:Carrier Proteins; D003300:Copper