Edman degradation

Before sequncing procss is initiated, it is necessary to break all non-covalent interation by denaturants (like high concentration of urea or GuHCl). This process will also separate subunits, incase of oligomeric proteins. Ocassionally, subunits of a oligomeric protein are connected by covalent interations. In that case special treatments are required to separate subunits. The protein is treated with Edman's reagent (phenyl isothiocyanate) which reacts with the N-terminal amino acid and under mild acidic condition forms a cyclic compound Phenyl thiohydantoin derivative (PTH−amino acid) of N-terminal amino acid is released. Amino acid of PTH −amino acid derivative is identified by chromatographic property of the PTH −amino acid derivative. In this process N-terminal aminos acid is identified after first cycle. Since this method proceeds from the N terminal residue, the reaction will not work if that N-terminal of a protein is blocked (generally due to post-translational modification) . After first cycle of the reaction, amino group of the second amino acid is free for reaction with Edman's reagent and at the end of reaction PTH derivative of second amino acid from N-terminal is released. The process continues till end of sequence or a disulfide bond is encountered in the sequence . PTH-cysteine derivative will remain attached with polypeptide and PTH-cystein will not be release (Fig. 1).