Escherichia coliSkp Chaperone Coexpression Improves Solubility and Phage Display of Single-Chain Antibody Fragments |
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Authors: | Andrew Hayhurst William J Harris |
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Institution: | Department of Molecular and Cell Biology, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, Scotland, AB25 2ZD |
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Abstract: | Expression of single-chain antibody fragments (scAb)in the periplasm ofEscherichia colioften results in low soluble product yield and cell lysis. We have increased scAb solubility and prevented cell culture lysis by coexpressing theE. coliSkp chaperone gene. A mutant Skp cistron was linked to a bacteriophage T7 gene 10 translational initiation region and placed either downstream of a scAb gene within an isopropyl β-
-thiogalactopyranoside-inducible expression cassette or on a separate colE1-compatible arabinose-inducible vector. Increases in scAb solubility reflected the amount of coexpressed Skp. A bacteriophage display vector that was also engineered to coexpress Skp permitted display of a virtually undisplayable scAb and should prove useful in expanding library sizes. |
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Keywords: | Skp chaperone single chain antibodies phage display dicistronic |
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