REDUCTIVE ALKYLATION OF AMINO GROUPS IN PROTEINS

When protein solutions are treated with low concentrations of simple aliphatic aldehydes or ketones and small amounts of sodium borohydride, amino groups are converted in high yield into the corresponding mono- or dialkylamino derivatives. Conditions for optimum reaction have been determined using gasliquid partition chromatography to follow the reductive alkylation of butylamine in aqueous solution. The reaction is strongly pH dependent. At pH 9.0 and 0°, only amino groups of proteins are modified. The principal product of the reductive methylation of proteins with formaldehyde was identified as ε-N,N-dimethyllysine; with acetaldehyde or acetone only the corresponding monoalkylated lysines were formed. The mild conditions and low concentrations of sodium borohydride used did not result in the reductive cleavage of disulfide bonds of lysozyme, insulin, ribonuclease, turkey ovomucoid, human serum transferrin, α-chymotrypsin, or chymotrypsinogen. Reductive methylation of ribonuclease produced an enzymatically inactive product with less than a single remaining unmodified lysine residue. Reductive alkylation of proteins occurs with minimal changes in gross physical properties as determined by studies of ultraviolet absorption spectra, sedimentation, and optical rotatory parameters. © 1968, American Chemical Society. All rights reserved.