High yield expression, refolding, and characterization of recombinant interferon alpha2/alpha8 hybrids in Escherichia coli.
Interferons (IFNs) are a family of pleiotropic cytokines used for the treatment of various viral infections and cancers. The low-cost production of IFNs with high biological value and the discovery of IFNs with improved properties are important for the treatment of these diseases as well as for understanding the physiological functions of these compounds. We describe a protein expression system for the production of IFNs alpha2, alpha8, and their hybrids in insoluble form in Escherichia coli, coupled to an efficient two-step optimized refolding and histidine-tag purification protocol. The expressed IFNs were of high biological value, as shown in antiviral and antiproliferative assays and some had specific activities higher than those of the commercially available interferon preparations and exhibited novel properties. This time-efficient, optimized protein expression method allows for the production of not just a single interferon subtype but several native and hybrid IFNs with relatively high yield and low cost that can be used in functional and potentially clinical assays.
Platis
Dimitris
D
Imperial College Faculty of Medicine at St. Mary's Hospital, South Wharf Road, London W2 1NY, UK.
Foster
Graham R
GR
D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D000998:Antiviral Agents / Q000494:pharmacology; D045744:Cell Line, Tumor; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D016898:Interferon-alpha / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D007372:Interferons / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010957:Plasmids; D015202:Protein Engineering; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment
D000970:Antineoplastic Agents; D000998:Antiviral Agents; C554164:IFNA8 protein, human; D016898:Interferon-alpha; D011993:Recombinant Fusion Proteins; D007372:Interferons