Selective cleavage of polypeptides with trifluoroacetic acid: applications for microsequencing

Cleavage of small polypeptides (less than 30 amino acid residues) by trifluoroacetic acid (TFA) under a variety of reaction conditions including time, temperature, TFA phase, and sample supports has been examined by N-terminal sequencing. Treatment with gas-phase TFA at room temperature will cleave polypeptide chains preferentially at the N-terminal side of serine and threonine residues. When liquid-phase TFA is used, additional cleavage at the C-terminal side of aspartic acid was detected. These procedures are applicable for directly treating samples immobilized on sequencer supports (glass fiber filters or polyvinylidene difluoride membranes) to verify the presence of a polypeptide with a blocked N-terminus as well as to obtain internal sequence data at subnanomole levels.