Date of Award
Doctor of Philosophy
Anti-DNA antibodies are hallmarks of SLE both in human and inbred lupus prone mice. The role of these antibodies in disease pathogenesis, particularly in glomerulonephritis has been documented. It is now well accepted that the genetic potential to generate anti-DNA antibodies also exists in the normal B cell immune repertoire. This genetic potential has been revealed by detection of anti-DNA antibodies in serum of normal individuals and by immortalizing DNA reactive B cells as hybridomas and as transformed cell lines. There is growing evidence that anti-DNA antibodies from the normal B cell repertoire and disease-associated anti-DNA antibodies share V region gene structures. Evidence for structural similarities includes the expression of normal monoclonal anti-DNA antibody idiotypes by lupus serum anti-DNA antibodies. Further direct evidence has been provided by sequence analysis of anti-DNA antibody genes. Thess studies showed that anti-DNA antibodies of either origin use no singular or simple mechanism for the generation of their specificity for DNA. It has been shown that VH and VL chain Ig germline genes in different combination can encode both normal and SLE anti-DNA antibodies and many of these genes can also be used by non DNA-binding antibodies. SLE anti-DNA antibodies are often somatically mutated and enriched with arginine residue(s) which could increase their affinity for DNA. In addition, the Ig V region of some anti-DNA antibodies are enriched with amino acids such as tyrosine, asparagine and glutamine which may also facilitate DNA-binding of these antibodies. It has also been suggested that heavy chains make a major contribution to DNA-binding in some anti-DNA antibodies based on the presence of common idiotype markers or motifs. One such motif YYGS, resides in the CDR3 region of KIM4.6 a natural human anti-DNA antibody heavy chain and is also expressed in more than 20% of human and murine anti-DNA antibodies and in the VH CDR2 region of some murine IgG anti-DNA antibodies.;The current study has explored the structural basis for DNA binding of human anti-DNA antibodies and in particular has focussed on the examination of the role of the heavy chain, the diversity region and the YYGS motif in conferring DNA specificity to the natural human monoclonal IgM anti-DNA antibody KIM4.6.;This study describes the generation and molecular characterization of Variants of KIM4.6 hybridoma which have lost their DNA binding property. The Ig V genes of three anti-Sm/RNP antibodies which either react with DNA or use genes related to KIM4.6 were also characterized. Gene manipulation techniques and a phage expression system or an in vitro transcription and translation system was used to further directly explore the role of YYGS motif in the DNA specificity of the KIM4.6 anti-DNA antibody.;This study revealed that: (1) the KIM4.6 heavy chain confers DNA specificity in this antibody, (2) the KIM4.6 D region YYGS motif is a necessary but not sufficient structural determinant in KIM4.6 reactivity to ds DNA but not ss DNA.
Mahmoudi, Mahmoud, "Structural Basis For Human Anti-dna Antibody Specificity" (1995). Digitized Theses. 2502.