Cobalt(2+) binding to human and tomato copper chaperone for superoxide dismutase: implications for the metal ion transfer mechanism.
The copper chaperone for superoxide dismutase (CCS) gene encodes a protein that is believed to deliver copper ions specifically to copper-zinc superoxide dismutase (CuZnSOD). CCS proteins from different organisms share high sequence homology and consist of three distinct domains; a CuZnSOD-like central domain 2 flanked by domains 1 and 3, which contain putative metal-binding motifs. We report deduced protein sequences from tomato and Arabidopsis, the first functional homologues of CCS identified in plants. We have purified recombinant human (hCCS) and tomato (tCCS) copper chaperone proteins, as well as a truncated version of tCCS containing only domains 2 and 3. Their cobalt(2+) binding properties in the presence and absence of mercury(2+) were characterized by UV-vis and circular dichroism spectroscopies and it was shown that hCCS has the ability to bind two spectroscopically distinct cobalt ions whereas tCCS binds only one. The cobalt binding site that is common to both hCCS and tCCS displayed spectroscopic characteristics of cobalt(2+) bound to four or three cysteine ligands. There are only four cysteine residues in tCCS, two in domain 1 and two in domain 3; all four are conserved in other CCS sequences including hCCS. Thus, an interaction between domain 1 and domain 3 is concluded, and it may be important in the copper chaperone mechanism of these proteins.
Zhu
H
H
Department of Chemistry and Biochemistry, University of California, Los Angeles, California 90095-1569, USA.
Shipp
E
E
Sanchez
R J
RJ
Liba
A
A
Stine
J E
JE
Hart
P J
PJ
Gralla
E B
EB
Nersissian
A M
AM
Valentine
J S
JS
D000595:Amino Acid Sequence; D017360:Arabidopsis; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003035:Cobalt / Q000737:chemistry*; D003545:Cysteine / Q000378:metabolism; D006801:Humans; D018551:Solanum lycopersicum; D008627:Mercuric Chloride / Q000494:pharmacology; D018832:Molecular Chaperones / Q000378:metabolism*; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000378:metabolism; D011485:Protein Binding; D016415:Sequence Alignment; D013053:Spectrophotometry; D013482:Superoxide Dismutase / Q000096:biosynthesis / Q000378:metabolism*
C114678:CCS protein, human; D018832:Molecular Chaperones; D010940:Plant Proteins; D003035:Cobalt; D008627:Mercuric Chloride; D013482:Superoxide Dismutase; D003545:Cysteine