Type VII collagen, the major component of anchoring fibrils, consists of a central collagenous triple-helical domain flanked by two noncollagenous domains, NC1 and NC2. The NC2 domain has been implicated in catalyzing the antiparallel dimer formation of
type VII procollagen. In this study, we produced the entire 161
amino acids of the NC2 domain plus 186
amino acids of adjacent collagenous domain (NC2/COL) and purified large quantities of the recombinant NC2/COL
protein. Recombinant NC2/COL readily formed
disulfide-bonded hexamers, each representing one antiparallel dimer of
collagen VII. Removal of the collagenous helical domain from NC2/COL by
collagenase digestion abolished the antiparallel dimer formation. Using site-directed mutagenesis, we found that mutation of either
cysteine 2802 or
cysteine 2804 alone within the NC2 domain blocked antiparallel dimer formation. In contrast, a single
cysteine mutation, 2634, within the collagenous helical domain had no effect. A generated
methionine to
lysine substitution, M2798K, that is associated with recessive
dystrophic epidermolysis bullosa, was unable to form antiparallel dimers. Furthermore,
autoantibodies from
epidermolysis bullosa acquisita patients also reacted with NC2/COL. We conclude that NC2 and its adjacent collagenous segment mediate antiparallel dimer formation of
collagen VII.
Epidermolysis bullosa acquisita autoantibodies bound to this domain may destabilize anchoring fibrils by interfering with antiparallel dimer assembly leading to epidermal-dermal disadherence.