ABSTRACT
A gene was chemically synthesised and expressed in Escherichia coli to produce [Ala30,32,33]IFN-alpha 2, an analogue of human alpha 2-interferon (IFN-alpha 2) which is devoid of activity on human cells. Eight additional analogues provided single changes in IFN-alpha 2 at each of these three conserved positions. No one residue is essential for activity, but both antiviral and anti-proliferative activity are particularly sensitive to changes in the side-chain of Arg33.
Subject(s)
Amino Acids/analysis , Interferon-gamma/physiology , DNA, Recombinant , Electrophoresis, Polyacrylamide Gel , Escherichia coli/genetics , Genes , Humans , Interferon-gamma/genetics , PlasmidsABSTRACT
A 511-base pair DNA fragment encoding human interferon-alpha 2 has been chemically synthesised and expressed from a lac UV5 and a synthetic trp promoter in Escherichia coli. The synthesis involved preparation of 68 oligodeoxyribonucleotides and their enzymic ligation. The product expressed from the trp promoter system had high antiviral activity and displayed biological effects similar to those of Namalwa interferon on natural killer cell activity and in a Daudi cell growth inhibition assay. E.coli minicells containing plasmid DNA with the synthetic IFN-alpha 2 gene under trp promoter control produce a protein with the same electrophoretic mobility as a sample of authentic IFN-alpha 2. The protein from E.coli cross-reacts with the monoclonal antibody NK-2 and was readily purified, close to homogeneity, by immunoadsorption chromatography on NK-2 sepharose.
