BACKGROUND: Autoantibodies directed against neutrophil proteinase 3 (PR3-ANCA) from patients with Wegener's granulomatosis and microscopic polyangiitis recognize conformational epitopes of PR3. During maturation of neutrophils, PR3 undergoes amino-terminal and carboxy-terminal processing. In contrast to amino-terminal processing, the effects of carboxy-terminal processing on recognition of PR3 by PR3-ANCA remain unknown. Carboxy-terminally modified or tagged recombinant PR3 (rPR3) molecules may be useful for the refinement of diagnostic assays and for the study of biological processes. METHODS: This study was designed to determine whether 293 cells can be used to express specifically designed carboxy-terminal variants of rPR3, and to evaluate the effects of different carboxy-terminal modifications on the recognition by PR3-ANCA in the capture ELISA. RESULTS: The rPR3-variants secreted into the media supernatants of transfected 293 cells escaped proteolytic processing. Furthermore, in contrast to the effects of amino-terminal pro-peptide deletion on PR3-ANCA binding, carboxy-terminal modifications (deletion and additions) did not significantly affect recognition by PR3-ANCA. CONCLUSIONS: This expression system is ideally suited for the expression of custom-designed carboxy-terminal rPR3 variants, and major conformational effects of carboxy-terminal modifications seem unlikely.
BACKGROUND: Autoantibodies directed against neutrophil proteinase 3 (PR3-ANCA) from patients with Wegener's granulomatosis and microscopic polyangiitis recognize conformational epitopes of PR3. During maturation of neutrophils, PR3 undergoes amino-terminal and carboxy-terminal processing. In contrast to amino-terminal processing, the effects of carboxy-terminal processing on recognition of PR3 by PR3-ANCA remain unknown. Carboxy-terminally modified or tagged recombinant PR3 (rPR3) molecules may be useful for the refinement of diagnostic assays and for the study of biological processes. METHODS: This study was designed to determine whether 293 cells can be used to express specifically designed carboxy-terminal variants of rPR3, and to evaluate the effects of different carboxy-terminal modifications on the recognition by PR3-ANCA in the capture ELISA. RESULTS: The rPR3-variants secreted into the media supernatants of transfected 293 cells escaped proteolytic processing. Furthermore, in contrast to the effects of amino-terminal pro-peptide deletion on PR3-ANCA binding, carboxy-terminal modifications (deletion and additions) did not significantly affect recognition by PR3-ANCA. CONCLUSIONS: This expression system is ideally suited for the expression of custom-designed carboxy-terminal rPR3 variants, and major conformational effects of carboxy-terminal modifications seem unlikely.
Authors: J M Clain; A M Hummel; J H Stone; F C Fervenza; G S Hoffman; C G M Kallenberg; C A Langford; W J McCune; P A Merkel; P A Monach; P Seo; R F Spiera; E W St Clair; S R Ytterberg; U Specks Journal: Clin Exp Immunol Date: 2017-02-07 Impact factor: 4.330
Authors: Lisa C Hinkofer; Susanne A I Seidel; Brice Korkmaz; Francisco Silva; Amber M Hummel; Dieter Braun; Dieter E Jenne; Ulrich Specks Journal: J Biol Chem Date: 2013-07-31 Impact factor: 5.157
Authors: J D Finkielman; P A Merkel; D Schroeder; G S Hoffman; R Spiera; E W St Clair; J C Davis; W J McCune; A Lears; S R Ytterberg; A M Hummel; M A Viss; T Peikert; J H Stone; U Specks Journal: Clin Exp Rheumatol Date: 2009 Jan-Feb Impact factor: 4.473
Authors: Yuan-Ping Pang; Marta Casal Moura; Gwen E Thompson; Darlene R Nelson; Amber M Hummel; Dieter E Jenne; Daniel Emerling; Wayne Volkmuth; William H Robinson; Ulrich Specks Journal: Front Immunol Date: 2019-10-25 Impact factor: 7.561
Authors: Alvise Berti; Sophie Hillion; Amber M Hummel; Young Min Son; Nedra Chriti; Tobias Peikert; Eva M Carmona; Wayel H Abdulahad; Peter Heeringa; Kristina M Harris; E William St Clair; Paul Brunetta; Fernando C Fervenza; Carol A Langford; Cees Gm Kallenberg; Peter A Merkel; Paul A Monach; Philip Seo; Robert F Spiera; John H Stone; Guido Grandi; Jie Sun; Jacques-Olivier Pers; Ulrich Specks; Divi Cornec Journal: JCI Insight Date: 2021-11-22