MANNAN-BINDING PROTEIN FORMS COMPLEXES WITH ALPHA-2-MACROGLOBULIN - A PROPOSED MODEL FOR THE INTERACTION

We report that alpha-2-macroglobulin (alpha(2)M) can form complexes with a high molecular weight porcine mannan-binding protein (pMBP-28). The alpha(2)M/pMBP-28 complexes were isolated by PEG-precipitation and affinity chromatography on mannan-Sepharose, protein A-Sepharose and anti-IgM Sepharose. The occurrence of alpha(2)M/pMBP-28 complexes was further indicated by crossed immunoelectrophoresis and by use of an anti-alpha(2)M affinity column and chelating Sepharose loaded with Zn2+. The eluates from these affinity columns showed alpha(2)M subunits (94 and 180 kDa) and pMBP subunits (28 kDa) in SDS-PAGE, which reacted with antibodies against alpha(2)M and pMBP-28, respectively, in Western blotting. Furthermore, the alpha(2)M/pMBP-28 complexes were demonstrated by electron microscopy. Fractionation of pMBP-containing D-mannose eluate from mannan-Sepharose on Superose 6 showed two protein peaks which reacted with anti-C1 s antibodies in ELISA, one of about 650-800 kDa, which in addition contained pMBP-28 and anti-alpha(2)M reactive material, the other with an M(r) of 100-150 kDa. The latter peak revealed rhomboid molecules (7 x 15 mn) in the electron microscope and a 67 kDa band in SDS-PAGE under reducing conditions. This band was also seen in eluates from the anti-alpha(2)M and chelating Sepharose columns. Based on these observations and previous findings by other investigators of a serine protease with about 67 kDa subunits which copurifies with human MBP we propose a model for the interaction of pMBP-28 with alpha(2)M.