Structural analysis of the heparin-binding site of the NC1 domain of collagen XIV by CD and NMR

Type XIV collagen, a fibril-associated collagen with interrupted triple hel ices (FACIT), interacts with the surrounding extracellular matrix and/or wi th cells via its binding to glycosaminoglycans (GAGs). To further character ize such interactions in the NC1 domain of chicken collagen XIV, we identif ied amino acids essential for heparin binding by affinity chromatography an alysis after proteolytic digestion of the synthetic peptide NC1(84-116). Th e 3D structure of this peptide was then obtained using circular dichroism a nd NMR. The NC1(84-116) peptide appeared poorly structured in water, but th e stabilization of its conformation by the interaction with hydrophobic sur faces or by using cosolvents (TFE, SDS) revealed a high propensity to adopt an alpha-helical folding. A 3D structure model of NC1(84-116), calculated from NMR data recorded in a TFE/water mixture, showed that the NC1-heparin binding site forms a amphipathic alpha-helix exhibiting a twisted basic gro ove. It is structurally similar to the consensus spatial alpha-helix model of heparin-binding [Margalit et al. (1993) J. Biol. Chem. 268, 19228-19231] , except that the GAG binding domain of NC1 may be extended over 18 residue s, that is, the NC1(94-111) segment. In addition, the formation of a hydrop hobic groove upon helix formation suggests the contribution of additional s equences to ensure the stability of the GAG-binding domain. Overall the NC1 (84-116) model exhibits a nativelike conformation which presents suitably o riented residues for the interaction with a specific GAG.