Initial work focused on the C-terminal tail residues of interferon only. Based on the computer design, two novel constructs, which we named RD1 and RD2, were prepared. In comparison with the IFN-alpha2c amino acid sequence, RD1 mutant contains only one amino acid change at position 163. RD2 mutant contains 8 amino acid changes. Both constructs were cloned into a yeast expression vector pKLAC2 and transformed into a host strainYap3 in which the YAP3 gene encoding yeast aspartic protease 3 (YAP3) was disrupted. Secreted IFN-alpha proteins were collected from culture supernatants and further purified using a reverse phase HPLC. Initial data showed that both RD1 and RD2 appear antiviral activities at a comparable level with the wild type IFN-alpha2a. Further characterization of purified RD1 and RD2 protein is ongoing.
| Tsuno, Takaya; Mejido, Josef; Zhao, Tongmao et al. (2009) IRF9 is a key factor for eliciting the antiproliferative activity of IFN-alpha. J Immunother 32:803-16 |
