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Epitope mapping of anti-glomerular basement membrane (GBM) antibodies with synthetic peptides

Hellmark, Thomas LU ; Brunmark, C; Trojnar, J and Wieslander, Jörgen LU (1996) In Clinical and Experimental Immunology 105(3). p.504-510
Abstract
Autoantibodies to the non-collagenous (NC1) domain of the alpha 3(IV)-chain of type IV collagen are found in sera from patients with anti-GBM nephritis. These antibodies have been shown to be pathogenic. In this study the antibody specificity has been investigated in patients with Goodpasture's syndrome and from a patient with atypical anti-GBM antibodies, recognizing the alpha 1(IV)-chain only. Overlapping synthetic peptides, covering the complete NC1 domains of the alpha 1(IV)- and alpha 3(IV)-chains were used in sandwich ELISA and competitive ELISA. None of the Goodpasture sera showed reactivity to the synthetic peptides. However, antibodies from the patient with atypical anti-GBM antibodies recognized a 20 amino acid peptide from the... (More)
Autoantibodies to the non-collagenous (NC1) domain of the alpha 3(IV)-chain of type IV collagen are found in sera from patients with anti-GBM nephritis. These antibodies have been shown to be pathogenic. In this study the antibody specificity has been investigated in patients with Goodpasture's syndrome and from a patient with atypical anti-GBM antibodies, recognizing the alpha 1(IV)-chain only. Overlapping synthetic peptides, covering the complete NC1 domains of the alpha 1(IV)- and alpha 3(IV)-chains were used in sandwich ELISA and competitive ELISA. None of the Goodpasture sera showed reactivity to the synthetic peptides. However, antibodies from the patient with atypical anti-GBM antibodies recognized a 20 amino acid peptide from the alpha 1(IV)-chain. The reactive peptide was further narrowed down with glycine substitution of the different amino acids. We have localized the epitope to the four last C-terminal amino acids of the alpha 1(IV)-chain, with the sequence 1754-MRRT. The two arginine residues were found to be essential for antibody binding. Threonine is important, while methionine is of less importance. These four amino acids are also determined to be the smallest peptide that could inhibit the binding of the autoantibodies to the native alpha 1(IV)-chain. This study shows that overlapping peptides can be used to map linear epitopes. However, for conformational epitopes such as the Goodpasture epitope, other methods must be used. It would be prognostically important to know the fine specificity of anti-GBM antibodies, since the patient with anti-alpha 1(IV) antibodies had a mild disease, while the Goodpasture patients with anti-alpha 3(IV) antibodies had a rapidly progressive disease. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
anti-GBM antibodies, Goodpasture syndrome, type IV collagen, synthetic peptides
in
Clinical and Experimental Immunology
volume
105
issue
3
pages
504 - 510
publisher
British Society for Immunology
external identifiers
  • pmid:8809141
ISSN
0009-9104
DOI
10.1046/j.1365-2249.1996.119808.x
language
English
LU publication?
yes
id
30930f5d-265f-491c-95df-cea85e222a02 (old id 1110059)
date added to LUP
2008-07-23 11:27:50
date last changed
2016-04-15 19:31:57
@article{30930f5d-265f-491c-95df-cea85e222a02,
  abstract     = {Autoantibodies to the non-collagenous (NC1) domain of the alpha 3(IV)-chain of type IV collagen are found in sera from patients with anti-GBM nephritis. These antibodies have been shown to be pathogenic. In this study the antibody specificity has been investigated in patients with Goodpasture's syndrome and from a patient with atypical anti-GBM antibodies, recognizing the alpha 1(IV)-chain only. Overlapping synthetic peptides, covering the complete NC1 domains of the alpha 1(IV)- and alpha 3(IV)-chains were used in sandwich ELISA and competitive ELISA. None of the Goodpasture sera showed reactivity to the synthetic peptides. However, antibodies from the patient with atypical anti-GBM antibodies recognized a 20 amino acid peptide from the alpha 1(IV)-chain. The reactive peptide was further narrowed down with glycine substitution of the different amino acids. We have localized the epitope to the four last C-terminal amino acids of the alpha 1(IV)-chain, with the sequence 1754-MRRT. The two arginine residues were found to be essential for antibody binding. Threonine is important, while methionine is of less importance. These four amino acids are also determined to be the smallest peptide that could inhibit the binding of the autoantibodies to the native alpha 1(IV)-chain. This study shows that overlapping peptides can be used to map linear epitopes. However, for conformational epitopes such as the Goodpasture epitope, other methods must be used. It would be prognostically important to know the fine specificity of anti-GBM antibodies, since the patient with anti-alpha 1(IV) antibodies had a mild disease, while the Goodpasture patients with anti-alpha 3(IV) antibodies had a rapidly progressive disease.},
  author       = {Hellmark, Thomas and Brunmark, C and Trojnar, J and Wieslander, Jörgen},
  issn         = {0009-9104},
  keyword      = {anti-GBM antibodies,Goodpasture syndrome,type IV collagen,synthetic peptides},
  language     = {eng},
  number       = {3},
  pages        = {504--510},
  publisher    = {British Society for Immunology},
  series       = {Clinical and Experimental Immunology},
  title        = {Epitope mapping of anti-glomerular basement membrane (GBM) antibodies with synthetic peptides},
  url          = {http://dx.doi.org/10.1046/j.1365-2249.1996.119808.x},
  volume       = {105},
  year         = {1996},
}