ABSTRACT
The number of each type of amino acid in a protein can be determined by acid hydrolysis and separation of the individual amino acids by ion exchange chromatography. The amino acids are detected by colorimetric reaction with, for example, ninhydrin or fluorescamine. In order to sequence an entire protein, the polypeptide chain has to be broken down into smaller fragments using either chemicals or enzymes. The amino-terminal residue of a protein can be identified by reacting the protein with a compound that forms a stable covalent link with the free a-amino group, prior to hydrolysis with 6 M HCl. The precise mass of intact proteins and peptides derived from them can be determined by mass spectrometry. Polypeptides can be chemically synthesized by covalently linking amino acids to the end of a growing polypeptide chain. In solid phase peptide synthesis the growing polypeptide chain is covalently anchored at its C-terminus to an insoluble support such as polystyrene beads.
