Isolation of Peptides for Microsequencing by In-Gel Proteolytic Digestion

Abstract

In contrast to predictions made at the advent of molecular biology about 15 years ago, the demand for classical amino acid sequence analysis has increased. However, the goal no longer is to determine the complete primary structure of a protein by direct sequencing, but rather to sequence a few relatively short peptides derived by proteolytic digestion of the purified protein. The sequences obtained are usually utilized for the design of oligonu-cleotide probes for subsequent cDNA cloning and DNA sequencing. Another important use of internal peptide sequences is the design and production of anti-peptide antibodies which have proved to be an extremely useful analytical and preparative tool. Yet another application is the positive identification of purified proteins.