Short peptide ligands for affinity purification of recombinant erythropoietin

M. M. MARANI; M. ETCHEVERRIGARAY; F. ALBERICIO; O. CASCONE; S. A. CAMPERI

Santa Fe

Congreso; XLII Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2006

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Recombinant erythropoietin (rhEPO) is used for therapeutics of anemia associated with chronic renal failure and for AZT-induced anemia of AIDS.  Monoclonal antibody (mAb) affinity chromatography and dye affinity chromatography are alternative techniques nowadays in use for its purification. mAb are expensive, thus increasing the cost of the final product. Affinity chromatography with Cibacron Blue as the ligand is widely used, but the selectivity is not high. The use of short peptides would result in a more economic process than with mAb and a more selective process than with dyes. Divide-couple-recombine (DCR) method allows obtaining a library with all possible combinations of the amino acids in the form of "one bead-one peptide". Peptide ligands can be selected from immunoaffinity screening of the library. A combinatorial library containing the tetrapeptides XXXX (X=all the natural amino acids except cysteine) was synthesised by the DAR method using Fmoc chemistry. The immunoaffinity screening of the solid-supported library for EPO was performed. Beads showing a positive reaction to rhEPO were isolated and peptide sequenced. Eight of the peptides were synthesised and immobilised on Sepharose. All the peptide ligands showed affinity for rhEPO. In future work the purification method will be optimised.