Purification and identification of recombinant hirudin and its degradation products expressed in Pichia pastoris

The purification and identification of recombinant hirudin (r‐hirudin) (rHV2‐Lys47) and its several C‐terminal proteolytic degradation derivatives, produced by Pichia pastoris, were described. The high‐purity rHV2‐Lys47 of above 99% and its three degradation products were obtained by a straightforward two‐step chromatography procedure, a combination of cation exchange and reverse phase chromatography, with a recovery yield of 74% for hirudin. The purified rHV2 had the predicted N‐terminal amino acid sequence and the derivatives were the degradation products of hirudin, short of one to three amino acid residues at C‐terminal.