University of Illinois Urbana-Champaign

Specificity and structure-function analysis of lupus-derived monoclonal anti-DNA autoantibodies

Rumbley, Catherine Ann

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https://hdl.handle.net/2142/20008

Description

Title
Specificity and structure-function analysis of lupus-derived monoclonal anti-DNA autoantibodies
Author(s)
Rumbley, Catherine Ann
Issue Date
1995
Doctoral Committee Chair(s)
Voss, Edward W., Jr.
Department of Study
Biology
Discipline
Biology
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
  • Health Sciences, Pathology
  • Health Sciences, Immunology
Language
eng
Abstract
  • Specificity and structure-function relationships of the monoclonal anti-DNA autoantibodies, BV 04-01 and BV 17-45, derived from the NZB/NZW F1 lupus-prone murine strain were examined. A single-chain Fv derivative of BV 04-01 was constructed and a series of site-specific mutants were engineered. A sensitive fluorescence-based tryptophan quenching assay was developed for analysis of the binding properties of the wild-type and mutant single-chain Fv proteins. Results of these binding assays revealed the importance of a tryptophan-thymidine-tyrosine complex in conveying ligand specificity in the cleft-like anti-DNA active site of BV 04-01. Basic residues within the active site enhanced the affinity of the interaction through energetically important but relatively non-specific contributions.
  • "The single-chain Fv studies were extended to the anti-dsDNA autoantibody BV 17-45. Various studies comparing the bivalent IgG molecule with the monovalent SCA derivative revealed a unique bispecific binding property of the BV 17-45 molecule. It was shown that the active site of BV 17-45 reacted with dsDNA and simultaneously displayed autoreactivity with its own Fc region as well as Fc regions of other antibody molecules of similar isotype. These two reactivities functioned cooperatively to form a stable, high affinity complex. A model was proposed which explained how such dual reactivities may qualify autoimmune antibodies as ""pathogenic"" for direct participation in such tissue destruction as observed in glomerulonephritis. The bispecific reactivity was also exhibited by BV 04-01 suggesting that this dual specificity may be a common feature of SLE-derived autoantibodies."
  • Finally, it was shown that BV 04-01 also displayed intrinsic catalytic activity for DNA molecules. The catalytic activity was exhibited by IgG, Fab, and SCA derivatives of BV 04-01 and was enhanced in the presence of divalent cations. Analysis of the catalytic properties of site-specific mutants led to the construction of a molecular model of the BV 04-01 catalytic active site.
Type of Resource
text
Permalink
http://hdl.handle.net/2142/20008
Copyright and License Information
Copyright 1995 Rumbley, Catherine Ann

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Graduate Theses and Dissertations at Illinois
Dissertations and Theses - Biology