Expression and purification of Canis interferon α in Escherichia coli using different tags

The potent and broad activity of Canis interferon a (CaIFN alpha) makes it an attractive candidate for the treatment of many viral diseases of dogs. Here, we fused CaIFN alpha to three different protein tags: thioredoxin (Trx), glutathione S-transferase (GST), and NusA (Nus), to facilitate its expression and purification in Escherichia coli. The Trx-CaIFN alpha and GST-CaIFN alpha fusion proteins formed inclusion bodies, while the Nus-CaIFN alpha, protein was soluble when expressed at low temperatures. Trx-CaIFN alpha was purified from inclusion bodies and refolded, while Nus-CaIFN alpha was purified under native conditions. The purity of Trx-CaIFN alpha and Nus-CaIFN alpha was greater than 90%, and their yields were 74.8% and 6.5%, respectively. Both Trx-CaIFN alpha and Nus-CaIFN alpha had antiviral activity in vitro. Their anti-viral activity was 1.09 +/- 0.47 x 10(14) and 2.25 +/- 0.87 x 10(12) U/mol, respectively, on Madin-Darby canine kidney cells. Both purification methods had advantages and disadvantages. A greater amount of Trx-CaIFN alpha was obtained, but refolding was required to obtain active protein. In contrast, soluble Nus-CaIFN alpha did not require refolding, which saved time and materials. However, Nus-CaIFN alpha, which contained a larger tag, had lower activity than Trx-CaIFN alpha. In general, we provided two protocols to obtain large amounts of CaIFN alpha with high antiviral activity. These protocols may promote the clinical development of CaIFN alpha in treating viral diseases in dog. (C) 2015 Elsevier Inc. All rights reserved.