Production of soluble single-chain T-cell receptor fragments in Escherichia coli trxB mutants

P. e. Molloy, W. J. Harris, G. Strachan, C. Watts, C. Cunningham (Lead / Corresponding author)

    Research output: Contribution to journalArticlepeer-review

    14 Citations (Scopus)


    Antibodies and T cell receptors (TCR) both belong to the immunoglobulin superfamily whose members are characterised by the possession of one or more immunoglobulin domains. The production of soluble single chain antibody fragments in Escherichia coli has, in recent years, become a routine laboratory procedure. In contrast, the production oft cell receptors in bacteria has remained problematic as the majority of the recombinant protein is insoluble. In this paper we show that single chain TCR produced in E. coli BL21 (DE3) and directed to the periplasm was also insoluble and that this was in part due to the failure of the cell protein processing machinery to cleave the pelB leader sequence. This problem was overcome by expressing the single chain TCR in the cytoplasm of E. coli which carry an inactive thioredoxin reductase gene. This strain allows the formation of disulphide bonds in the cell cytoplasm which we believe encourages the correct folding of the recombinant protein. We have constructed both a human and mouse single chain TCR in these bacteria and demonstrated using BIAcore technology that these molecules have folded in a conformation which allows their recognition by conformational specific ligands. In addition, we have used one of our soluble single chain TCR preparations to isolate a TCR specific Fab molecule from a phage antibody library.

    Original languageEnglish
    Pages (from-to)73-81
    Number of pages9
    JournalMolecular Immunology
    Issue number2
    Publication statusPublished - 15 Feb 1998


    • Phage antibody
    • Single chain T cell receptor
    • T cell receptors
    • Thioredoxin reductase

    ASJC Scopus subject areas

    • Immunology
    • Molecular Biology


    Dive into the research topics of 'Production of soluble single-chain T-cell receptor fragments in Escherichia coli trxB mutants'. Together they form a unique fingerprint.

    Cite this