Hydrophobic Amino Acid Residues Are Critical for the Immunodominant Epitope of the Goodpasture Autoantigen

• Published in: The Journal of biological chemistry Vol. 276; no. 9; pp. 6370 - 6377
• Main Authors: David, Michelle, Borza, Dorin-Bogdan, Leinonen, Anu, Belmont, John M., Hudson, Billy G.
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 02.03.2001; American Society for Biochemistry and Molecular Biology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M008956200

Goodpasture (GP) autoimmune disease is caused by autoantibodies to type IV collagen that bind to the glomerular basement membrane, causing rapidly progressing glomerulonephritis. The immunodominant GPAautoepitope is encompassed by residues 17–31 (the EAregion) within the noncollagenous (NC1) domain of the α3(IV) chain. The GP epitope is cryptic in the NC1 hexamer complex that occurs in the type IV collagen network found in tissues and inaccessible to autoantibodies unless the hexamer dissociates. In contrast, the epitope for the Mab3 monoclonal antibody is also located within the EA region, but is fully accessible in the hexamer complex. In this study, the identity of residues that compose the GPA autoepitope was determined, and the molecular basis of its cryptic nature was explored. This was achieved using site-directed mutagenesis to exchange the α3(IV) residues in the EAregion with the corresponding residues of the homologous but non-immunoreactive α1(IV) NC1 domain and then comparing the reactivity of the mutated chimeras with GPA and Mab3 antibodies. It was shown that three hydrophobic residues (Ala18, Ile19, and Val27) and Pro28 are critical for the GPA autoepitope, whereas two hydrophilic residues (Ser21 and Ser31) along with Pro28 are critical for the Mab3 epitope. These results suggest that the cryptic nature of the GPA autoepitope is the result of quaternary interactions of the α3, α4, and α5 NC1 domains of the hexamer complex that bury the one or more hydrophobic residues. These findings provide critical information for understanding the etiology and pathogenesis of the disease as well as for designing drugs that would mimic the epitope and thus block the binding of GP autoantibodies to autoantigen.