Characterization of T cell receptor single-chain Fv fragments secreted by myeloma cells.
Myeloma cells have been used to produce milligram quantities of soluble alpha beta T cell receptor (TCR) molecules as single-chain polypeptides in which the TCR variable (V) domains are connected by a peptide linker (TCR scFv). Unlike most TCR scFv produced in bacteria, the purified TCR scFv were stable and showed no tendency to aggregate when kept at concentrations up to 10 mg/ml. Circular dichroism analyses of the TCR scFv indicated that they contained a high proportion of beta-pleated sheet structures. Since the V alpha subunits present in the TCR scFv contained their own signal sequences, they provided the opportunity to determine by N-terminal amino acid sequencing the position of the signal cleavage of three distinct mouse V alpha. Two of the experimentally determined signal cleavage sites differed from those previously predicted on the basis of biochemical and statistical criteria. The expression approach outlined in this report has been applicable to three distinct alpha beta TCR and should contribute to the large scale production of soluble TCR amenable to structural studies.
Centre d'Immunologie INSERM-CNRS de Marseille-Luminy, France.
C
C
Grégoire
B
B
Malissen
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G
Mazza
D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D000939:Epitopes; D006825:Hybridomas; D051379:Mice; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000737:chemistry / Q000235:genetics*; D011993:Recombinant Fusion Proteins; D012995:Solubility; D013601:T-Lymphocytes / Q000276:immunology*
D000939:Epitopes; D016693:Receptors, Antigen, T-Cell, alpha-beta; D011993:Recombinant Fusion Proteins