A new semi-empirical method for the determination of the subunit molecular weight of a protein

We have developed a semi-empirical method to determine the subunit molecular weight of a protein. The method is a minor modification of the Edman degradation and is based on a simple chemical procedure to modify specifically the N-terminal NH2 group. Initially, all NH2 groups, including the epsilon-NH2 groups of lysine residues and the N-terminal alpha-NH2 group, are reacted with phenylisothiocyanate and the protein derivative is subjected to one step of the Edman degradation. The newly exposed N terminus is then reacted with radioactivity labelled phenylisothiocyanate. A value for the subunit molecular weight can be obtained from an analysis of the incorporated radioactivity and the amount of the protein in the sample. The molecular weight of five different proteins have been determined by this method. Our method is particularly useful for proteins containing lipid or sugar components and also for relatively small peptides. The procedure described in this paper for the specific modification of the N terminus has been found to be a powerful tool for protein sequencing.