SP-A binding sites on bovine alveolar macrophages

Surfactant protein A (SP-A) binding to bovine alveolar macrophages was exam ined in order to characterize SP-A binding proteins on the cell surface and to isolate putative receptors from these cells that could be obtained in l arge amounts. Human SP-A, unlabeled or labeled with gold particles, was bou nd to freshly isolated macrophages and analyzed with ELISA or the transmiss ion electron microscope. Binding of SP-A was inhibited by Ca2+ chelation, b y an excess of unlabeled SP-A, or by the presence of 20 mg/ml mannan, We co nclude that bovine alveolar macrophages expose binding sites for SP-A that are specific and that depend on Ca2+ and on mannose residues. For isolation of SP-A receptors with homologous SP-A as ligand we isolated SP-A from bov ine lung lavage. SDS-PAGE analysis of the purified SP-A showed a protein of 32-36 kDa, Functional integrity of the protein was demonstrated. Bovine SP -A bound to Dynabeads was used to isolate SP-A binding proteins. From the f ractionated and blotted proteins of the receptor preparation two proteins b ound SP-A in a Ca2+-dependent manner, a 40-kDa protein showing mannose depe ndency and a 210-kDa protein, showing no mannose sensitivity, (C) 1998 Acad emic Press.