Expression of recombinant Lym-1 single-chain Fv in Escherichia coli.
Lym-1 single-chain Fv (sFv) can be used for targeted radiodiagnosis and therapy of B-lymphocytic malignancies. Lym-1 sFv was constructed and expressed as a glutathione S-transferase fusion protein, using a (G4S)3 linker connecting the C-terminus of the VH domain and the N-terminus of the VL domain of Lym-1. Six histidine residues and an E Tag epitope were introduced at the C-terminus of the sFv. Lym-1 sFv was purified with glutathione-Sepharose 4B affinity chromatography followed by digestion with thrombin. Lym-1 sFv of 28 kDa was confirmed by Western blotting with anti-(E Tag) monoclonal antibody. An antigen binding assay of Lym-1 and a CD study indicated that it is functionally active.
Department of Food Science and Technology, Chungnam National University, Taejon 305-764, Korea.
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Bin Song
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Won
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Meares
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology; D000954:Antigens, Surface / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology*; D001483:Base Sequence; D002846:Chromatography, Affinity / Q000379:methods; D002942:Circular Dichroism; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism*; D015870:Gene Expression / Q000235:genetics; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology*; D007945:Leukemia, Lymphoid / Q000276:immunology / Q000628:therapy; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins