The goodpasture autoantigen -: Identification of multiple cryptic epitopes on the NC1 domain of the α3(IV) collagen chain

Goodpasture (GP) disease is an autoimmune disorder in which autoantibodies against the alpha 3(TV) chain of type TV collagen bind to the glomerular and alveolar basement membranes, causing progressive glomerulonephritis and pulmonary hemorrhage, Two major conformational epitope regions have been identified on the noncollagenous domain of type IV collagen (NC1 domain) of the alpha 3(IV) chain as residues 17-31 (E-A) and 127-141 (E-B) (Netzer, K.-O. et al. (1999) J. Biol. Chem. 274, 11267-11274), To determine whether these regions are two distinct epitopes or form a single epitope, three GP sera were fractionated by affinity chromatography on immobilized NC1 chimeras containing the E-A and/or the E-B region. Four subpopulations of GP antibodies with distinct epitope specificity for the alpha 3(IV)NC1 domain were thus separated and characterized. They were designated GP(A), GP(B), GP(AB) and GP(X), to reflect their reactivity with E-A only, E-B only, both regions, and neither, respectively. Hence, regions E-A and E-B encompass critical amino acids that constitute three distinct epitopes for GP(A), GP(B), and GP(AB) antibodies, respectively, whereas the epitope for GP, antibodies is located in a different unknown region. The GP(A) antibodies were consistently immunodominant, accounting for 60-65% of the total immunoreactivity to alpha 3(IV)NC1; thus, they probably play a major role in pathogenesis. Regions E-A and E-B are held in close proximity because they jointly form the epitope for Mab3, a monoclonal antibody that competes for binding with GP autoantibodies. All GP epitopes are sequestered in the hexamer configuration of the NC1 domain found in tissues and are inaccessible for antibody binding unless dissociation of the hexamer occurs, suggesting a possible mechanism for etiology of GP disease. GP antibodies have the capacity to extract alpha 3(IV)NC1 monomers, but not dimers, from native human glomerular basement membrane hexamers, a property that may be of fundamental importance for the pathogenesis of the disease.