Reactions of Glycine-containing Peptides with Cupric Ions and with p-Nitrophenyl Acetate

Abstract

The reactivity of groups in proteins may be studied in terms of their rates of reaction to form stable or unstable products or in terms of equilibria with small molecules or ions (2). The equilibria between cupric ions and peptides are of particular interest because of the stability and spectral properties of the complexes formed and the biological importance of the copper proteins. It has been found not only that Cu(I1) ion combines with the terminal amino group in dipeptides but that it may combine with the amide nitrogen at the same time with displacement of the proton from the amide nitrogen. The latter step is complete for dipeptides by pH 7, and very stable structures are formed (1, 3-6). Potentially, therefore, the NH2 terminus of a peptide chain in a protein may be a very strong binding site for Cu(I1) ion, probably surpassing the individual imidazole group or amino group (7). Titrations of mixtures of Cu(I1) ions with diglycylglycine and triglycylglycine have also indicated that the process of formation of Cu(II)-N bonds with displacement of the amide proton may continue through a second and even third peptide linkage, making possible an even more elaborate and stable complex between Cu(I1) and the first 3 or 4 residues in a peptide chain (8). The present report confirms the titration studies under the conditions of temperature and ionic strength used previously for the dipeptide studies and confirms directly the role of the second amide N atom by showing the effect of substitution of sarcosine residues in glycylglycylsarcosine and glycylsarcosylglycine. The ability of a Cu(II)-dipeptide complex bearing an additional hydroxyl ion coordinated to the Cu(I1) to catalyze the hydrolysis of p-nitrophenyl acetate (1) has prompted the use of this catalytic test to confirm the displacement of amide protons instead of the additional coordination of hydroxyl ions in the Cu(II)-tripeptide and tetrapeptide systems. Last, the characteristics of the visible absorption spectra of the various complexes are tabulated, and a band in the ultraviolet region is described.