Abstract Sperm whale ferrimyoglobin was treated with 0.2 m sodium bromoacetate at pH 6.8 to test the accessibility for alkylation of certain residues, particularly those of histidine. After a plateau had been reached in the reaction process, the product was shown to retain native properties. Electrophoretically, it was free of unmodified material and contained four main components apparently differing by unit gradations of net charge. The most fully studied preparation contained 5.3, 4.6, 1.3, 0.9, and 17.9 residues per molecule of histidine, 1,3-dicarboxymethylhistidine, 3-carboxymethylhistidine, 1-carboxymethylhistidine, and lysine, respectively. The methionine content was not affected. The aminoterminal valyl residue was largely alkylated. Assignment of histidyl residues according to the derivatives identified in peptide fragments was made according to the method of the preceding paper in the series (Hugli, T. E., and Gurd, F. R. N., J. Biol. Chem., 245, 1930 (1970)). The following residues were recovered as unmodified histidine: 24, 36, 82, 93, and 97. The other residues were isolated in various states of alkylation. The results establish a considerable number of points of similarity between the reactivity patterns in solution and in the crystalline state. Most points of contrast with the results of alkylation in the crystalline states can be ascribed to the absence of neighboring lattice molecules. However, the fact that residue 36 becomes unreactive when the protein passes from the crystalline to the dissolved state implies a distinct structural change controlling the accessibility of the residue to the alkylating agent.