Variable region cDNA sequences and antigen binding specificity of mouse monoclonal antibodies to isomaltosyl oligosaccharides coupled to proteins. T-dependent analogues of alpha(1----6)dextran.
Four mouse hybridomas specific for alpha(1----6)dextran, 16.4.12E (IgA kappa, C57BL/6), 28.4.10A (IgM kappa, BALB/c), 35.8.2H (IgG1 kappa, BALB/c), and 36.1.2D (IgM kappa, BALB/c) were obtained by immunization with the T-dependent Ag isomaltohexaose or isomaltotriose coupled to keyhole limpet hemocyanin or to BSA. Immunochemical characterization of the hybridoma antibodies showed that 16.4.12E and 36.1.2D had cavity-type combining sites, recognizing the terminal non-reducing end of alpha(1----6)dextran, whereas 28.4.10A and 35.8.2H had groove-type sites, recognizing internal linear segments of the dextran. The V region cDNA of the H and L chains of the antibodies were cloned and sequenced. VH of 16.4.12E and VH of 36.1.2D belonged to the X24 and Q52 germ-line gene families, respectively. The VH and V kappa sequences of 16.4.12E and V kappa sequence of 36.1.2D were highly homologous to those of W3129, the only anti-alpha(1----6)dextran mAb with a cavity-type site thus far sequenced; 16.4.12E differed from W3129 in the D, JH, and J kappa. VH genes of 28.4.10A and 35.8.2H were homologous to those of several anti-alpha(1----6)dextrans with groove-type sites, but belonged to the J558 germ-line gene family, differed from the other J558 anti-alpha(1----6)dextrans, probably representing a different germ-line subfamily. The L chain sequence of 28.4.10A encoded by V kappa-Ars and J kappa 2 was almost identical to other groove-type anti-alpha(1----6)dextrans obtained by immunizing with the T-independent glycolipid Ag, stearyl-isomaltotetraose. Use of T-dependent Ag such as isomaltosyl oligosaccharide-protein conjugates provides an additional parameter for probing the fine structure of antibody combining sites and evaluating the V-gene repertoire of anti-alpha(1----6)dextrans.
Matsuda
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T
Department of Microbiology, College of Physicians and Surgeons, Columbia University, New York 10032.
Kabat
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EA
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis; D000918:Antibody Specificity; D001483:Base Sequence; D001666:Binding Sites, Antibody; D002352:Carrier Proteins / Q000276:immunology; D004247:DNA / Q000302:isolation & purification; D003911:Dextrans / Q000276:immunology*; D004187:Disaccharides / Q000276:immunology*; D006825:Hybridomas / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D007145:Immunoglobulin kappa-Chains / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007534:Isomaltose / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D013601:T-Lymphocytes / Q000276:immunology*
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D003911:Dextrans; D004187:Disaccharides; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D007145:Immunoglobulin kappa-Chains; C041999:alpha-1,6-dextran; D007534:Isomaltose; D004247:DNA