Glomerular basement membrane: Identification of dimeric subunits of the noncollagenous domain (hexamer) of collagen IV and the goodpasture antigen

The noncollagenous (NC1) domain hexamer of glomerular basement membrane (GBM) collagen is composed of a multiplicity of monomeric and dimeric subunits, and specific subunits are the targets for anti-GBM autoantibodies of patients with Goodpasture (GP) syndrome. The identity of GBM monomers has been established and the α3(IV)NC1 monomer identified as the one that binds GP antibodies (Gunwar, S., Saus, J., Noelken, M. E., and Hudson, B.G. (1990) J. Biol. Chem. 265, 5466-5469). In the present study, the chain origin of 25 dimeric components and the identity of those that bound the anti-GBM antibodies from two GP patients were determined. This was accomplished by NH₂-terminal sequence analysis and immunoblotting analysis of dimeric components that were resolved by two-dimensional electrophoresis in combination with high pressure liquid chromatography. The results revealed that (a) the components are mainly homodimers of the NC1 domains of α1, α2, α3, α4, and probably α5 chains of collagen IV, reflecting a specificity of protomer-protomer association and (6) each homodimer had several size and charge isoforms. The GP antibodies bound exclusively to both α3(IV)NC1 monomers and dimers and not to other basement membrane constituents. These findings provided new insights about the structure of GBM collagen and together with our previous findings firmly established the α3(IV) chain as the target for the anti-GBM antibodies that mediate glomerulonephritis and pulmonary hemorrhage in patients with Goodpasture syndrome.