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. 1995 Oct;102(1):112–119. doi: 10.1111/j.1365-2249.1995.tb06644.x

Epitope mapping of anti-proteinase 3 and anti-myeloperoxidase antibodies.

L Chang 1, S Binos 1, J Savige 1
PMCID: PMC1553353  PMID: 7554377

Abstract

Anti-proteinase 3 (PR3) and anti-myeloperoxidase (MPO) autoantibodies are present in many patients with Wegener's granulomatosis (WG) and microscopic polyarteritis. The aim of this study was to determine whether these antibodies bound to linear peptide sequences on their target antigens. If common linear epitopes were demonstrated, then these could be manufactured and used in diagnostic ELISAs for anti-PR3 and anti-MPO antibodies. In addition, any homology between these epitopes and bacterial or viral sequences might implicate those microorganisms in the development of these antibodies and the pathogenesis of the associated diseases. The presence of linear epitopes on PR3 and MPO was suggested by the binding of the corresponding autoantibodies to these proteins after they had been reduced with beta-mercaptoethanol (beta-ME) and denatured with SDS or boiling, and digested with proteases. Four of the 22 sera with anti-PR3 antibodies bound to PR3 in Western blots after treatment with SDS, beta-ME and boiling for 5 min. Thermal denaturation reduced the amount of binding more than other forms of denaturation. One serum with anti-PR3 antibodies bound to Lys-C and Glu-C-digested PR3 in dot blots. Linear epitopes could not be further defined by their binding in an ELISA using overlapping peptides corresponding to the PR3 molecule because of non-specific binding. Three of the five sera with anti-MPO antibodies bound to MPO in Western blots after treatment with SDS, beta-ME and boiling for 5 min. One serum with anti-MPO antibodies bound to Lys-C and Glu-C-digested MPO in dot blots. Again, linear epitopes could not be further defined using an ELISA with overlapping peptides because of non-specific binding. Some anti-PR3 and anti-MPO antibodies are likely to recognize linear epitopes, but these cannot be defined by use of a PIN ELISA system.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barakat S., Briand J. P., Abuaf N., van Regenmortel M. H., Muller S. Mapping of epitopes on U1 snRNP polypeptide A with synthetic peptides and autoimmune sera. Clin Exp Immunol. 1991 Oct;86(1):71–78. doi: 10.1111/j.1365-2249.1991.tb05776.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bini P., Gabay J. E., Teitel A., Melchior M., Zhou J. L., Elkon K. B. Antineutrophil cytoplasmic autoantibodies in Wegener's granulomatosis recognize conformational epitope(s) on proteinase 3. J Immunol. 1992 Aug 15;149(4):1409–1415. [PubMed] [Google Scholar]
  3. Brocke S., Brautbar C., Steinman L., Abramsky O., Rothbard J., Neumann D., Fuchs S., Mozes E. In vitro proliferative responses and antibody titers specific to human acetylcholine receptor synthetic peptides in patients with myasthenia gravis and relation to HLA class II genes. J Clin Invest. 1988 Dec;82(6):1894–1900. doi: 10.1172/JCI113807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Davies D. J., Moran J. E., Niall J. F., Ryan G. B. Segmental necrotising glomerulonephritis with antineutrophil antibody: possible arbovirus aetiology? 1982 Aug 28-Sep 4Br Med J (Clin Res Ed) 285(6342):606–606. doi: 10.1136/bmj.285.6342.606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Elkon K. B. Use of synthetic peptides for the detection and quantification of autoantibodies. Mol Biol Rep. 1992 Jun;16(3):207–212. doi: 10.1007/BF00464709. [DOI] [PubMed] [Google Scholar]
  6. Ewing C., Ebringer R., Tribbick G., Geysen H. M. Antibody activity in ankylosing spondylitis sera to two sites on HLA B27.1 at the MHC groove region (within sequence 65-85), and to a Klebsiella pneumoniae nitrogenase reductase peptide (within sequence 181-199). J Exp Med. 1990 May 1;171(5):1635–1647. doi: 10.1084/jem.171.5.1635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Falk R. J., Becker M., Terrell R., Jennette J. C. Anti-myeloperoxidase autoantibodies react with native but not denatured myeloperoxidase. Clin Exp Immunol. 1992 Aug;89(2):274–278. doi: 10.1111/j.1365-2249.1992.tb06944.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Falk R. J., Jennette J. C. Anti-neutrophil cytoplasmic autoantibodies with specificity for myeloperoxidase in patients with systemic vasculitis and idiopathic necrotizing and crescentic glomerulonephritis. N Engl J Med. 1988 Jun 23;318(25):1651–1657. doi: 10.1056/NEJM198806233182504. [DOI] [PubMed] [Google Scholar]
  9. Geysen H. M., Rodda S. J., Mason T. J., Tribbick G., Schoofs P. G. Strategies for epitope analysis using peptide synthesis. J Immunol Methods. 1987 Sep 24;102(2):259–274. doi: 10.1016/0022-1759(87)90085-8. [DOI] [PubMed] [Google Scholar]
  10. Hines J. J., Danho W., Elkon K. B. Detection and quantification of human anti-Sm antibodies using synthetic peptide and recombinant SmB antigens. Arthritis Rheum. 1991 May;34(5):572–579. doi: 10.1002/art.1780340509. [DOI] [PubMed] [Google Scholar]
  11. Kao R. C., Wehner N. G., Skubitz K. M., Gray B. H., Hoidal J. R. Proteinase 3. A distinct human polymorphonuclear leukocyte proteinase that produces emphysema in hamsters. J Clin Invest. 1988 Dec;82(6):1963–1973. doi: 10.1172/JCI113816. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Mori T., Sugawa H., Piraphatdist T., Inoue D., Enomoto T., Imura H. A synthetic oligopeptide derived from human thyrotropin receptor sequence binds to Graves' immunoglobulin and inhibits thyroid stimulating antibody activity but lacks interactions with TSH. Biochem Biophys Res Commun. 1991 Jul 15;178(1):165–172. doi: 10.1016/0006-291x(91)91794-d. [DOI] [PubMed] [Google Scholar]
  13. Niles J. L., McCluskey R. T., Ahmad M. F., Arnaout M. A. Wegener's granulomatosis autoantigen is a novel neutrophil serine proteinase. Blood. 1989 Nov 1;74(6):1888–1893. [PubMed] [Google Scholar]
  14. Oldstone M. B. Molecular mimicry and autoimmune disease. Cell. 1987 Sep 11;50(6):819–820. doi: 10.1016/0092-8674(87)90507-1. [DOI] [PubMed] [Google Scholar]
  15. Scofield R. H., Harley J. B. Autoantigenicity of Ro/SSA antigen is related to a nucleocapsid protein of vesicular stomatitis virus. Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3343–3347. doi: 10.1073/pnas.88.8.3343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Snook J. A., Chapman R. W., Fleming K., Jewell D. P. Anti-neutrophil nuclear antibody in ulcerative colitis, Crohn's disease and primary sclerosing cholangitis. Clin Exp Immunol. 1989 Apr;76(1):30–33. [PMC free article] [PubMed] [Google Scholar]
  17. Tuaillon N., Muller S., Pasquali J. L., Bordigoni P., Youinou P., Van Regenmortel M. H. Antibodies from patients with rheumatoid arthritis and juvenile chronic arthritis analyzed with core histone synthetic peptides. Int Arch Allergy Appl Immunol. 1990;91(3):297–305. doi: 10.1159/000235131. [DOI] [PubMed] [Google Scholar]
  18. Williams R. C., Jr, Staud R., Malone C. C., Payabyab J., Byres L., Underwood D. Epitopes on proteinase-3 recognized by antibodies from patients with Wegener's granulomatosis. J Immunol. 1994 May 1;152(9):4722–4737. [PubMed] [Google Scholar]
  19. de Vries D. D., Dekker-Saeys A. J., Gyodi E., Bohm U., Ivanyi P. Absence of autoantibodies to peptides shared by HLA-B27.5 and Klebsiella pneumoniae nitrogenase in serum samples from HLA-B27 positive patients with ankylosing spondylitis and Reiter's syndrome. Ann Rheum Dis. 1992 Jun;51(6):783–789. doi: 10.1136/ard.51.6.783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. van de Wiel B. A., Dolman K. M., van der Meer-Gerritsen C. H., Hack C. E., von dem Borne A. E., Goldschmeding R. Interference of Wegener's granulomatosis autoantibodies with neutrophil Proteinase 3 activity. Clin Exp Immunol. 1992 Dec;90(3):409–414. doi: 10.1111/j.1365-2249.1992.tb05860.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. van der Woude F. J., Rasmussen N., Lobatto S., Wiik A., Permin H., van Es L. A., van der Giessen M., van der Hem G. K., The T. H. Autoantibodies against neutrophils and monocytes: tool for diagnosis and marker of disease activity in Wegener's granulomatosis. Lancet. 1985 Feb 23;1(8426):425–429. doi: 10.1016/s0140-6736(85)91147-x. [DOI] [PubMed] [Google Scholar]

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