STRUCTURE, FUNCTION AND TISSUE FORMS OF THE C-TERMINAL GLOBULAR DOMAIN OF COLLAGEN-XVIII CONTAINING THE ANGIOGENESIS INHIBITOR ENDOSTATIN

The C-terminal domain NC1 of mouse collagen XVIII (38 kDa) and the sho rter mouse and human endostatins (22 kDa) were prepared in recombinant form from transfected mammalian cells. The NC1 domain aggregated non- covalently into a globular trimer which was partially cleaved by endog enous proteolysis into several monomers (25-32 kDa) related to endosta tin. Endostatins were obtained in a highly soluble, monomeric form and showed a single N-terminal sequence which, together with other data, indicated a compact folding. Endostatins and NC1 showed a comparable b inding activity for the microfibrillar fibulin-1 and fibulin-2, and fo r heparin. Domain NC1, however, was a distinctly stronger ligand than endostatin for sulfatides and the basement membrane proteins laminin-l and perlecan. Immunological assays demonstrated endostatin epitopes o n several tissue components (22-38 kDa) and in serum (120-300 ng/ml), the latter representing the smaller variants. The data indicated that the NC1 domain consists of an N-terminal association region (similar t o 50 residues), a central protease-sensitive hinge region (similar to 70 residues) and a C-terminal stable endostatin domain (similar to 180 residues). They also demonstrated that proteolytic release of endosta tin can occur through several pathways, which may lead to a switch fro m a matrix-associated to a more soluble endocrine form.