Characterisation of a novel cysteine/histidine-rich metal binding domain from Xenopus nuclear factor XNF7.
A 42 amino acid synthetic peptide corresponding to a newly defined cysteine/histidine-rich protein motif called B-box, from the Xenopus protein XNF7 has been characterised. The metal-binding stoichiometry and dissociation constant for zinc were determined by competition with the chromophoric chelator Br2BAPTA, demonstrating that one zinc atom binds per molecule of peptide despite the presence of seven putative metal ligands, and represents the first application of this method to measuring zinc stoichiometry of proteins and/or peptides. Cobalt binding studies indicate that the motif binds zinc more tightly than cobalt, that cysteines are used as ligands and that the cation is co-ordinated tetrahedrally. Circular dichroism and NMR studies both indicate that the B-box peptide is structured only in the presence of zinc, copper and to a lesser extent cobalt.
Borden
K L
KL
National Institute for Medical Research, The Ridgeway, London, UK.
Martin
S R
SR
O'Reilly
N J
NJ
Lally
J M
JM
Reddy
B A
BA
Etkin
L D
LD
Freemont
P S
PS
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002614:Chelating Agents; D002942:Circular Dichroism; D003545:Cysteine / Q000032:analysis*; D004268:DNA-Binding Proteins; D004527:Egg Proteins; D004533:Egtazic Acid / Q000031:analogs & derivatives; D006639:Histidine / Q000032:analysis*; D009682:Magnetic Resonance Spectroscopy; D008670:Metals / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D014981:Xenopus; D029867:Xenopus Proteins
D002614:Chelating Agents; D004268:DNA-Binding Proteins; D004527:Egg Proteins; D008670:Metals; D009687:Nuclear Proteins; D010750:Phosphoproteins; D029867:Xenopus Proteins; C053459:5,5'-dibromo-1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; C071551:XNF7 protein, Xenopus; D006639:Histidine; D004533:Egtazic Acid; D003545:Cysteine