THE ROLE OF THE AMINO-TERMINAL DOMAIN AND THE COLLAGENOUS REGION IN THE STRUCTURE AND THE FUNCTION OF RAT SURFACTANT PROTEIN-D

Surfactant protein D (SP-D) is a member of the C-type lectin superfami ly with four distinct structural domains: an amino terminus involved i n forming intermolecular disulfides, a collagen-like domain, a neck re gion, and a carbohydrate recognition domain, A collagen domain deletio n mutant (CDM) of SP-D was created by site-directed mutagenesis, A sec ond variant lacking both the amino-terminal region and the collagen-li ke domain was generated by collagenase treatment and purification of t he collagenase-resistant fragment (CRF), The CDM expressed in CHO-K1 c ells formed the covalent trimers, but not the noncovalent dodecamers, typical of native SP-D, The CRF derived from recombinant SP-D formed o nly monomers. The CDM bound mannose-Sepharose and phosphatidylinositol (PI) as well as SP-D, but the binding to mannosyl bovine serum albumi n and glucosylceramide was diminished by approximately 60%. The CRF di splayed weak binding to mannose-Sepharose and PI and essentially no bi nding to mannosyl bovine serum albumin and glucosylceramide. Both SP-D and CDM altered the self-aggregation of PI-containing liposomes. SP-D reduced the density and the light scattering properties of PI aggrega tes. These results demonstrate that the collagen-like domain is requir ed for dodecamer but not covalent trimer formation of SP-D and plays a n important, but not essential, role in the interaction of SP-D with P I and GlcCer. Removal of the amino-terminal domain of SP-D along with the collagen-like domain diminishes PI binding and effectively elimina tes GlcCer binding.