Design, engineering and production of functional single-chain T cell receptor ligands

• Published in: Protein engineering 1999-09, Vol.12 (9), p.771-778
• Main Authors: Burrows, G.G., Chang, J.W., Bächinger, H-P., Bourdette, D.N., Offner, H., Vandenbark, A.A.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Base Sequence; DNA; drug design; Escherichia coli; Histocompatibility Antigens Class II - chemistry; Histocompatibility Antigens Class II - metabolism; Humans; immunotherapy; Ligands; MHC class II; Molecular Sequence Data; Protein Engineering; Rats; Receptors, Antigen, T-Cell - metabolism; Sequence Homology, Amino Acid; TCR ligand; β1α1
• ISSN: 0269-2139; 1741-0126; 1460-213X; 1741-0134
• DOI: 10.1093/protein/12.9.771

Major histocompatibility complex (MHC) class II molecules are membrane-anchored heterodimers on the surface of antigen presenting cells (APCs) that bind the T cell receptor, initiating a cascade of interactions that results in antigen-specific activation of clonal populations of T cells. The peptide binding/T cell recognition domains of rat MHC class II (alpha-1 and beta-1 domains) were expressed as a single exon for structural and functional characterization. These recombinant single-chain T cell receptor ligands (termed `β1α1' molecules) of approximately 200 amino acid residues were designed using the structural backbone of MHC class II molecules as template, and have been produced in Escherichia coli with and without N-terminal extensions containing antigenic peptides. Structural characterization using circular dichroism predicted that these molecules retained the antiparallel β-sheet platform and antiparallel α-helices observed in the native MHC class II heterodimer. The proteins exhibited a cooperative two-state thermal folding–unfolding transition. β1α1 molecules with a covalently linked MBP-72–89 peptide showed increased stability to thermal unfolding relative to the empty β1α1 molecules. This new class of small soluble polypeptide provides a template for designing and refining human homologues useful in detecting and regulating pathogenic T cells.