Goodpasture syndrome. Localization of the epitope for the autoantibodies to the carboxyl-terminal region of the alpha 3(IV) chain of basement membrane collagen.
The autoantibodies of patients with Goodpasture syndrome are primarily targeted to the noncollagenous (NC1) domain of the alpha 3(IV) chain of basement membrane collagen (Saus, J., Wieslander, J., Langeveld, J. P. M., Quinones, S., and Hudson, B. G. (1988) J. Biol. Chem. 263, 13374-13380). In the present study, the location of the Goodpasture epitope in human alpha 3NC1 was determined, and its structure was partially characterized. This was achieved by identification of regions of alpha 3NC1 which are candidates for the epitope and which are structurally unique among the five known homologous NC1 domains (alpha 1-alpha 5); amino acids that are critical for Goodpasture antibody binding, by selective chemical modifications; and regions that are critical for Goodpasture antibody binding, by synthesis of 12 alpha 3NC1 peptides and measurement of their antibody binding capacity. The carboxyl-terminal region, residues 198-233, was identified as the most likely region for the epitope. By experiment, lysine and cysteine were identified as critical amino acids for antibody binding. Three synthetic peptides were found to inhibit Goodpasture antibody binding to alpha 3NC1 markedly: a 36-mer (residues 198-233), a 12-mer (residues 222-233), and a 5-mer (residues 229-233). Together, these results strongly indicate that the Goodpasture epitope is localized to the carboxyl-terminal region of alpha 3NC1, encompassing residues 198-233 as the primary antibody interaction site and that its structure is discontinuous. These findings provide a conceptual framework for future studies to elucidate a more complete epitope structure by sequential replacement of residues encompassing the epitope using cDNA expression products and peptides synthesized chemically.
Kalluri
R
R
Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City 66103.
Gunwar
S
S
Reeders
S T
ST
Morrison
K C
KC
Mariyama
M
M
Ebner
K E
KE
Noelken
M E
ME
Hudson
B G
BG
D000595:Amino Acid Sequence; D000818:Animals; D019867:Anti-Glomerular Basement Membrane Disease / Q000276:immunology*; D001323:Autoantibodies / Q000032:analysis*; D001485:Basement Membrane / Q000276:immunology / Q000502:physiology; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D002417:Cattle; D003094:Collagen / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D006801:Humans; D046911:Macromolecular Substances; D008961:Models, Structural; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000276:immunology; D011487:Protein Conformation; D012689:Sequence Homology, Nucleic Acid
D001323:Autoantibodies; D000939:Epitopes; D046911:Macromolecular Substances; D010455:Peptides; D003094:Collagen