A proportion of proteinase 3 (PR3)-specific anti-neutrophil cytoplasmic antibodies (ANCA) only react with PR3 after cleavage of its N-terminal activation dipeptide

J. Sun, D. N. Fass, M. A. Viss, A. M. Hummel, H. Tang, H. A. Homburger, U. Specks

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35 Scopus citations


ANCA directed against PR3 are highly specific for Wegener's granulomatosis and microscopic polyangiitis, and have been implicated in the pathogenesis of small vessel vasculitis. Most PR3-ANCA are directed against conformational epitopes on PR3. This study was designed to determine whether the cleavage of the N-terminal activation dipeptide of PR3 is required for the binding of PR3-ANCA. Recombinant PR3 (rPR3) variants were expressed in the epithelial cell line, 293. As confirmed by radiosequencing, the rPR3 secreted into the 293 cell culture supernatant is N-terminally unprocessed. Two enzymatically inactive rPR3 mutants were expressed in 293 cells: rPR3-S176A and Δ-rPR3-S176A. rPR3-S176A contains the N-propetide Ala-2-Glu-1, Δ-rPR3-S176A does not. Culture supernatants of rPR3- S176A and Δ-rPR3-S176A expressing 293 cells were used as sources of target antigen for PR3-ANCA testing by capture ELISA. Forty unselected consecutive PR3-ANCA+ sera were tested. With Δ-rPR3- S176A as antigen all 40 were recognized, compared with only 34 of 40 when rPR3-S176A served as target antigen. The majority of the serum samples contained a mixture of antibodies reacting with epitopes accessible on the mature and on the proform of PR3. In conclusion, the cleavage of the N-terminal activation dipeptide of PR3 is not an absolute requirement for recognition by all PR3-ANCA. However, a substantial proportion of PR3-ANCA recognize (a) target antigen(s) exposed only after the conformational change of PR3 associated with the N-terminal processing. In 15% of sera this PR3-ANCA subset occurred exclusively. PR3-ANCA subtypes can be differentiated using specifically designed rPR3 variants as target antigens, and non- haematopoietic mammalian cells without regulated secretory pathway can be used for their expression.

Original languageEnglish (US)
Pages (from-to)320-326
Number of pages7
JournalClinical and Experimental Immunology
Issue number2
StatePublished - Nov 17 1998



  • Anti-neutrophil
  • Cytoplasmic antibodies
  • N-terminal propeptide
  • Recombinant proteinase 3
  • Target antigen

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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