Expression of soluble and active interferon consensus in SUMO fusion expression system in E. coli

Peciak, Karolina; Tommasi, Rita; Choi, Ji-Won; Brocchini, Steve; Laurine, Emmanuelle

doi:10.1016/j.pep.2014.03.009

Cited by 25 publications

(33 citation statements)

References 49 publications

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“…The results showed that the Nus tag increased the solubility of CaIFNα (Nus-CaIFNα) efficiently, which means that the larger tag may help the production of soluble proteins (Fig.1). This is in accordance with many other studies, which showed that the co-expression of a target protein with a highly soluble partner facilitates its soluble expression [25,27]. CaIFNα with tags fused at its N-terminus did not lose antiviral activity.…”

Section: Discussionsupporting

confidence: 92%

“…Additionally, the purification process was relatively complicated. Compared with other expression systems, E. coli expression systems have favorable attributes for protein production, such as rapid expression, high expression yields, low costs, and simple scale-up processes [25]. The most significant advantage of an E. coli expression system is that it produces target protein rapidly, only requiring about 12 h. There are many successful examples in which interferon with high anti-viral activity were expressed in E. coli.…”

Section: Discussionmentioning

confidence: 99%

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

Yang

Pan

Chen

et al. 2015

Protein Expression and Purification

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Section: Discussionsupporting

confidence: 92%

Section: Discussionmentioning

confidence: 99%

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

Yang

Pan

Chen

et al. 2015

Protein Expression and Purification

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“…Previous studies have also demonstrated that different types of recombinant interferons produced by SUMO fusion technology showed increased antiviral or antiproliferative activity (Peciak et al . ; Tileva et al . ).…”

Section: Discussionmentioning

confidence: 99%

“…For instance, Peciak et al . () have reported the successful expression of interferon consensus protein (IFN‐con) using a combination of SUMO‐tagged construct and SHuffle ® strain. Based on their results, 86–88% of total expressed protein was recovered in soluble fraction.…”

Section: Discussionmentioning

confidence: 99%

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Cytosolic expression of functional Fab fragments in Escherichia coli using a novel combination of dual SUMO expression cassette and EnBase^® cultivation mode

et al. 2017

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A Generic Protocol for Purifying Disulfide-Bonded Domains and Random Protein Fragments Using Fusion Proteins with SUMO3 and Cleavage by SenP2 Protease

Besir¹

2017

Methods in Molecular Biology

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Recombinant expression of heterologous proteins in E. coli is well established for a wide range of proteins, although in many cases, purifying soluble and properly folded proteins remains challenging (Sorensen and Mortensen, J Biotechnol 115:113-128, 2005; Correa and Oppezzo, Methods Mol Biol 1258:27-44, 2015). Proteins that contain disulfide bonds (e.g., cytokines, growth factors) are often particularly difficult to purify in soluble form and still need optimizing of protocols in almost every step of the process (Berkmen, Protein Expr Purif 82:240-251, 2012; de Marco, Microb Cell Fact 11:129, 2012). Expression of disulfide bonded proteins in the periplasm of E. coli is one approach that can help to obtain soluble protein with the correct disulfide bridges forming in the periplasm. This offers the appropriate conditions for disulfide formation although periplasmic expression can also result in low expression levels and incorrect folding of the target protein (Schlapschy and Skerra, Methods Mol Biol 705:211-224, 2011). Generation of specific antibodies often requires a specific antigenic sequence of a protein in order to get an efficient immune response and minimize cross-reactivity of antibodies. Larger proteins like GST (Glutathione-S-transferase) or MBP (maltose binding protein) as solubilizing fusion partners are frequently used to keep antigens soluble and immunize animals. This approach has the disadvantage that the immune response against the fusion partner leads to additional antibodies that need to be separated from the antigen-specific antibodies. For both classes of proteins mentioned above, a protocol has been developed and optimized using the human version of small ubiquitin-like modifier 3 (SUMO3) protein and its corresponding protease SenP2. This chapter describes the experimental steps for expression, purification, refolding, and cleavage that are applicable to both disulfide-bonded proteins with a defined structure and random protein fragments for antibody generation or larger peptides with defined sequence that are difficult express on their own.

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Expression of soluble and active interferon consensus in SUMO fusion expression system in E. coli

Cited by 25 publications

References 49 publications

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

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

Cytosolic expression of functional Fab fragments in Escherichia coli using a novel combination of dual SUMO expression cassette and EnBase^® cultivation mode

A Generic Protocol for Purifying Disulfide-Bonded Domains and Random Protein Fragments Using Fusion Proteins with SUMO3 and Cleavage by SenP2 Protease

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Expression of soluble and active interferon consensus in SUMO fusion expression system in E. coli

Cited by 25 publications

References 49 publications

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

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

Cytosolic expression of functional Fab fragments in Escherichia coli using a novel combination of dual SUMO expression cassette and EnBase® cultivation mode

A Generic Protocol for Purifying Disulfide-Bonded Domains and Random Protein Fragments Using Fusion Proteins with SUMO3 and Cleavage by SenP2 Protease

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Cytosolic expression of functional Fab fragments in Escherichia coli using a novel combination of dual SUMO expression cassette and EnBase^® cultivation mode