Design, engineering, and production of human recombinant t cell receptor ligands derived from human leukocyte antigen DR2.
Major histocompatibility complex (MHC) class II molecules are membrane-anchored heterodimers on the surface of antigen-presenting cells that bind the T cell receptor, initiating a cascade of interactions that results in antigen-specific activation of clonal populations of T cells. Susceptibility to multiple sclerosis is associated with certain MHC class II haplotypes, including human leukocyte antigen (HLA) DR2. Two DRB chains, DRB5*0101 and DRB1*1501, are co-expressed in the HLA-DR2 haplotype, resulting in the formation of two functional cell surface heterodimers, HLA-DR2a (DRA*0101, DRB5*0101) and HLA-DR2b (DRA*0101, DRB1*1501). Both isotypes can present an immunodominant peptide of myelin basic protein (MBP-(84-102)) to MBP-specific T cells from multiple sclerosis patients. We have previously demonstrated that the peptide binding/T cell recognition domains of rat MHC class II (alpha1 and beta1 domains) could be expressed as a single exon for structural and functional characterization; Burrows, G. G., Chang, J. W., Bächinger, H.-P., Bourdette, D. N., Wegmann, K. W., Offner, H., and Vandenbark A. A. (1999) Protein Eng. 12, 771-778; Burrows, G. G., Adlard, K. L., Bebo, B. F., Jr., Chang, J. W., Tenditnyy, K., Vandenbark, A. A., and Offner, H. (2000) J. Immunol. 164, 6366-6371). Single-chain human recombinant T cell receptor ligands (RTLs) of approximately 200 amino acid residues derived from HLA-DR2b were designed using the same principles and have been produced in Escherichia coli with and without amino-terminal extensions containing antigenic peptides. Structural characterization using circular dichroism predicted that these molecules retained the antiparallel beta-sheet platform and antiparallel alpha-helices observed in the native HLA-DR2 heterodimer. The proteins exhibited a cooperative two-state thermal unfolding transition, and DR2-derived RTLs with a covalently linked MBP peptide (MBP-(85-99)) showed increased stability to thermal unfolding relative to the empty DR2-derived RTLs. These novel molecules represent a new class of small soluble ligands for modulating the behavior of T cells and provide a platform technology for developing potent and selective human diagnostic and therapeutic agents for treatment of autoimmune disease.
Chang
J W
JW
Department of Neurology, Shriner's Hospital for Children, and Department of Biochemistry and Molecular Biology, Oregon Health Sciences University, Portland, Oregon 97201, USA.
Mechling
D E
DE
Bächinger
H P
HP
Burrows
G G
GG
D000595:Amino Acid Sequence; D001483:Base Sequence; D006684:HLA-DR Antigens / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D015802:HLA-DR2 Antigen / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D004676:Myelin Basic Protein / Q000235:genetics; D010446:Peptide Fragments / Q000235:genetics; D015202:Protein Engineering; D017433:Protein Structure, Secondary; D011948:Receptors, Antigen, T-Cell / Q000819:agonists*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013816:Thermodynamics
D006684:HLA-DR Antigens; D015802:HLA-DR2 Antigen; D008024:Ligands; D004676:Myelin Basic Protein; D010446:Peptide Fragments; C432412:RTL303 protein, human; D011948:Receptors, Antigen, T-Cell; D011994:Recombinant Proteins; C107908:myelin basic protein 85-99