Properties of whole human IgG molecules produced by the expression of cloned anti-DNA antibody cDNA in mammalian cells.
Antibodies to DNA are believed to play an important role in systemic lupus erythematosus (SLE). High affinity IgG antibodies which show marked specificity for double stranded DNA (dsDNA) are particularly closely linked to the occurrence and severity of tissue damage. Sequence analysis of mouse and human monoclonal antibodies has previously suggested that mutations in the complementarity determining regions (CDRs) play a major role in determining these binding properties. In many cases such mutations increase the overall number of basic residues in the CDRs. To further elucidate the role played by such mutations it is important to develop methods of expressing cloned autoantibody cDNA in the form of functional whole immunoglobulin molecules. We describe a system in which autoantibody VH and VL cDNA from monoclonal human anti-DNA antibodies, B3 and WRI176 were cloned into separate vectors which allowed their expression as whole heavy and whole light chains respectively. By cotransfecting mammalian cells with pairs of heavy and light chain vectors it was possible to produce whole IgG molecules from each of the four possible VH/VL combinations. Only antibody produced by homologous VH and VL pairs bound DNA, suggesting that in these autoantibodies both chains are important in conferring this property.
Rahman
M A
MA
Bloomsbury Rheumatology Unit/Division of Rheumatology, Department of Medicine, University College, London, UK.
Kettleborough
C A
CA
Latchman
D S
DS
Isenberg
D A
DA
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D001323:Autoantibodies / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D019556:COS Cells; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D003851:Deoxyribonucleases / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D008180:Lupus Erythematosus, Systemic / Q000276:immunology; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D018076:DNA, Complementary; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D003851:Deoxyribonucleases