Binding of mouse and human fibulin-2 to extracellular matrix ligands.

Recombinant mouse and human fibulin-2 were obtained as disulfide-bonded trimers from transfected kidney cell clones and used in solid phase, biosensor and radioligand binding assays. Strong binding occurred with fibronectin and required calcium. A distinct interaction was also observed with nidogen but this was only partially blocked by EDTA. Distinctly weaker affinities were detected for collagen IV, perlecan and the N-terminal globule of collagen VI alpha 3 chain, while no or only little binding activity could be detected for several other collagen types, laminin-1, BM-40, fibulin-1 and vitronectin. This weaker binding reactions were also dependent on calcium. Surface plasmon resonance assays demonstrated for fibulin-2 binding to nidogen and fibronectin high equilibrium dissociation constants (0.5 to 1 microM) due to a rapid initial dissociation of the complexes. This is apparently followed by a slower stabilizing reaction. The fibulin-2 binding site of nidogen could be localized to its C-terminal globular domain G3, which also possesses a high-affinity binding site for laminin-1. Several tests demonstrated competition between the two ligands, probably due to steric hindrance. Binding of nidogen to immobilized fibulin-2 allowed the formation of ternary complexes with collagen IV, perlecan and fibulin-1, which, as shown previously, bind independently of the G3 domain. This indicated multifunctional binding properties for fibulin-2 and several alternative routes for its integration into basement membranes and other extracellular structures.