Expression And Purification Of Recombinant Canine Interferon-a Using Self-Cleaving SUMO/ULP1System

SUMO consists of about100amino acids, a highly conserved peptide family found in eukaryotes. As a fusion tag, SUMO can increase expression level and solubility of foreign proteins. The SUMO tag can be cleaved from the fusion protein by SUMO protease ULP1. Elastin-like polypeptide (ELP) is a class of synthetic molecules with VPGXG repeats with temperature-induced reversible transition property. Therefore, the expression system based on ELP and SUMO/ULP1can be used to express and purify recombinant proteins.To very the usability of the self-cleaving SUMO/ULP1system for expression and purification of recombinant proteins, the coding sequences for SUMO and ULP1were amplified from yeast genome by PCR and submitted to sequence analysis. The SUMO and ULP1sequences were inserted into prokaryotic expression vector containing ELP sequence, resulting in fusion expression vectors pELP-SUMO and pET-ELP-ULPl. The coding sequence for canine interferon-a was amplified by PCR and subcloned into pELP-SUMO vector, resulting in fusion expression vector pET-ELP-SUMO-CaIFN. After IPTG induction at low temperature, SDS-PAGE detected an expected74-kDa protein band in the lysate of pET-ELP-SUMO-CalFN-transformed E.coli, with a yield of about15-18mg/L. By using the temperature-induced transition property of ELP, the fusion protein was purified after repeated centrifugation at different temperatures. By co-incubation of fusion protein ELP-SUMO-CaIFN with fusion protein ELP-ULP1, recombinant canine interferon-a was successfully released. These data indicate the correctness of cloned SUMO and ULP1sequences and the established expression system can be used to express and purify recombinant proteins.