Sharma, Arti; (2002) Analysis of structure and function of anti-dsDNA antibodies. Doctoral thesis (Ph.D.), University College London (United Kingdom).

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Abstract

Anti-dsDNA antibodies occur spontaneously in the serum of patients with systemic lupus erythematosus (SLE). The appearance of serum anti-dsDNA antibodies correlates with nephritis both in mice and human patients. In prior studies, a dominant peptide motif (D/EWD/EYS/G) that reacts with R4A, a pathogenic mouse monoclonal anti-DNA antibody was identified using a phage display library. The synthetic R4A peptide DWEYS inhibited the dsDNA binding of R4A and its deposition in the kidney of SCID mice. I have investigated the binding of human polyclonal and monoclonal anti-dsDNA and anti-cardiolipin antibodies to a peptide mimotope of dsDNA (R4A peptide) and screened random peptide libraries with human polyclonal affinity purified anti-dsDNA antibodies. I have also studied the crossreactivity of human monoclonal and polyclonal anti-dsDNA antibodies from sera of SLE patients to the antigen phosphorylcholine (PC) and investigated in humans, the relationship between anti-PC antibodies and anti-dsDNA antibodies expressing the 8.12 light chain idiotype, by using Fabs derived from a phage display combinatorial library. The major results obtained were as follows: DNA binding by four IgG and two IgM anti-DNA monoclonal antibodies and cardiolipin binding of four IgM monoclonal anti-cardiolipin antibodies was inhibited by the R4A peptide. Polyclonal anti-dsDNA and anti-cardiolipin antibodies were found to be inhibited by a multimeric form of the peptide. No consensus peptide motif was obtained on screening phage display libraries with affinity purified human anti-dsDNA antibodies. However, the YHDWDYY motif (shown in bold) was identified representing a part of a previously identified consensus motif (D/EWD/EYS/G) for R4A. The binding of polyclonal anti-DNA antibodies to dsDNA was inhibited by PC though reciprocal studies of inhibition of PC binding by DNA was not observed. This would suggest that only a fraction of anti-PC antibodies crossreact with dsDNA. The results obtained suggest that Fabs expressing the 8.12 idiotype and showing dsDNA and/or PC binding use VH3 gene family for their heavy chains and Vλ3, Vλ4 and Vλ6 gene families for their light chains.

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