Reversible disruption by cupric ions of myoglobin and its carboxamidomethyl derivative.

Carboxamidomethyl derivatives of sperm whale metmyoglobin were prepared by reaction at pH 6.8 with iodoacetamide. The modified protein is similar to previously studied carboxymethyl preparations and shows many of the properties of the native, unmodified protein. Preparations were obtained with compositions in the following ranges, expressed as moles of histidine derivative per mole of protein: histidine 4.8 to 7.5 ; dicarboxamidomethylhistidine, 1.6 to 4.6; l-carboxamidomethylhistidine, trace to 0.5; and d-carboxamidomethylhistidine, 1.6 to 2.6. Lysine was converted to the extent of 0.5 to 2.4 residues into the dicarboxamidomethyl derivative, out of a total of 19 lysine residues. The cupric complexes of the modified preparations are relatively soluble and easily studied in terms of maximum binding and of absorption and optical rotatory dispersion spectra near pH 6. The maximum binding of cupric ions was reduced by about the degree of conversion of histidine residues to the dicarboxamidomethyl derivative. The modified proteins were indistinguishable from the unmodified protein with respect to changes in absorption spectrum and optical rotatory dispersion in the Soret region upon copper (II) binding, and the changes were essentially fully reversible by EDTA. The modified proteins underwent smaller, reversible changes in rotation at 233 rnp on the addition of cupric ions. The inability of a dicarboxamidomethylhistidine derivative to react with cupric ion near pH 6 was conlirmed with a derivative prepared from acetylglycylglycyl-L-histidine. The results are interpreted as indicating that the carboxamidomethyl modification reaction has little if any effect on the ability of cupric ions to disrupt the heme-protein relationship, but greatly alters the ability of cupric ions to bring about a second step in disruption of the protein structure.