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Expressivity tag: a novel tool for increased expression in Escherichia coli

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avis › Tidsskriftartikel › Forskning › peer review

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  • Jon Gade Hansted, Danmark
  • Laura Pietikäinen
    • ,
  • Friederike Hög
    • ,
  • Hans Uffe Sperling-Petersen, Danmark
  • Kim Kusk Mortensen
Protein expression in Escherichia coli is rarely trivial as low expression and insolubility are common problems. In this work we define a fusion partner, which increases expression levels similarly to the distinct function of solubility and affinity tags. This type of fusion tag we term an expressivity tag. Our work is based on earlier observations where 3' deletions of the InfB gene displays strongly increased expression levels. We have constructed progressively shortened fragments of the InfB(1-471) gene and fused gene fragments to a gfp reporter gene. A 5-fold increase in GFP expression was seen for an optimal 21 nucleotide InfB(1-21) sequence compared to gfp independently. We defined the InfB(1-21) sequence as an expressivity tag. The tag was tested for improved expression of two biotechnological important proteins streptavidin and a single chain antibody (scFv). Expression of both streptavidin and scFv(L32) was improved as evaluated by SDS-PAGE. Calculation of folding energies in the translation initiation region gave higher free energies for gfp, L32 and streptavidin when linked to InfB(1-21) than independently. InfB(1-21) did however not improve the codon usage or codon adaptation index. The expressivity tag is an important addition to the box of tools available for optimizing heterologous protein expression.
OriginalsprogEngelsk
TidsskriftJournal of Biotechnology
Vol/bind155
Nummer3
Sider (fra-til)275-83
Antal sider9
ISSN0168-1656
DOI
StatusUdgivet - 2011

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