NC1 domain of type VII collagen binds to the beta3 chain of laminin 5 via a unique subdomain within the fibronectin-like repeats.
Type VII collagen, the major component of anchoring fibrils, consists of a central collagenous triple-helical domain flanked by two noncollagenous, globular domains, NC1 and NC2. Approximately 50% of the molecular mass of the molecule is consumed by the NC1 domain. We previously demonstrated that NC1 binds to various extracellular matrix components including a complex of laminin 5 and laminin 6 (Chen et al, 1997a). In this study, we examined the interaction of NC1 with laminin 5 (a component of anchoring filaments). Both authentic and purified recombinant NC1 bound to human and rat laminin 5 as measured by enzyme-linked immunosorbant assay and by binding of 125I-radiolabeled NC1 to laminin 5-coated wells, but not to laminin 1 or albumin. NC1 bound predominantly to the beta3 chain of laminin 5, but also to the gamma2 chain when examined by a protein overlay assay. The binding of 125I-NC1 to laminin 5 was inhibited by a 50-fold excess of unlabeled NC1 or de-glycosylated NC1, as well as a polyclonal antibody to laminin 5 or a monoclonal antibody to the beta3 chain. In contrast, the NC1-laminin 5 interaction was not affected by a monoclonal antibody to the alpha3 chain. Using NC1 deletion mutant recombinant proteins, a 285 AA (residues 760-1045) subdomain of NC1 was identified as the binding site for laminin 5. IgG from an epidermolysis bullosa acquisita serum containing autoantibodies to epitopes within NC1 that colocalized with the laminin 5 binding site inhibited the binding of NC1 to laminin 5. Thus, perturbation of the NC1-laminin 5 interaction may contribute to the pathogenesis of epidermolysis bullosa acquisita.
Chen
M
M
Department of Dermatology, Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Marinkovich
M P
MP
Jones
J C
JC
O'Toole
E A
EA
Li
Y Y
YY
Woodley
D T
DT
D000818:Animals; D003094:Collagen / Q000737:chemistry / Q000378:metabolism*; D016107:Epidermolysis Bullosa Acquisita / Q000097:blood; D005353:Fibronectins / Q000737:chemistry*; D006801:Humans; D007797:Laminin / Q000378:metabolism*; D051379:Mice; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism