Sites of Binding of Copper(II) Ion by Peptide (1–24) of Bovine Serum Albumin

• Published in: The Journal of biological chemistry 1968-07, Vol.243 (14), p.3817-3825
• Main Authors: Bradshaw, R A, Shearer, W T, Gurd, F R
• Format: Article
• Language: English
• Subjects: Alkylation; Amino Acid Sequence; Amino Acids - analysis; Animals; Cattle; Chelating Agents; Chemical Phenomena; Chemistry, Physical; Chromatography, Ion Exchange; Copper; Histidine; Peptides; Serum Albumin, Bovine; Spectrophotometry; Spectrum Analysis
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(18)92017-x

1. The amino-terminal peptide (1–24) of bovine serum albumin was subjected to carboxymethylation by 0.2 m bromoacetate at pH 6.8 in the presence of 0, 1, and 2 eq of copper(II) ion. The treatment with bromoacetate in the absence of copper(II) caused almost quantitative carboxymethylation of the imidazole groups of the 3 histidyl residues at positions 3, 9, and 18 in the sequences. In addition, the terminal α-amino group was apparently modified. Spectral measurements on the copper-containing complexes showed little or no effect on the copper chelation by the alkylation treatment. 2. Binding of 1 eq of copper(II) protects quantitatively the terminal α-amino group and histidyl residue 3, and appears to reduce somewhat the reactivity of histidyl residue 18. 3. Binding of 2 eq of copper(II) also protects the components of the preferred terminal site, but, in addition, both histidyl residues 9 and 18 are largely, and about equally, protected from alkylation. The binding of the 2nd eq therefore appears to involve both histidyl residues, in keeping with spectral and titration results. An intramolecular chelate structure or intermolecular chelates involving dimers or higher polymers are suggested. 4. The tetrapeptide l -aspartyl- l -threonyl- l -histidyl- l -lysine, which corresponds to the first 4 residues of peptide (1–24), was shown by titration, absorbance, and circular dichroism measurements to form an equimolar complex with copper(II) that has substantially the properties of the complex of peptide (1–24) containing 1 eq of copper(II) ion. Similar measurements on the equimolar complex of bovine serum albumin fill out a continuity that is especially evident from measurements of circular dichroism.