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Mannan‐Binding Protein Forms Complexes with α‐2‐Macroglobulin. A Proposed Model for the Interaction
Author(s) -
STORGAARD P.,
NIELSEN E. HOLM,
SKRIVER E.,
ANDERSEN O.,
SVEHAG S.E.
Publication year - 1995
Publication title -
scandinavian journal of immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.934
H-Index - 88
eISSN - 1365-3083
pISSN - 0300-9475
DOI - 10.1111/j.1365-3083.1995.tb03670.x
Subject(s) - sepharose , alpha 2 macroglobulin , chemistry , affinity chromatography , mannan , fractionation , alpha (finance) , microbiology and biotechnology , protein subunit , biochemistry , chromatography , macroglobulin , biology , enzyme , polysaccharide , medicine , gene , construct validity , nursing , patient satisfaction
We report that α‐2‐macroglobulin (α 2 M) can form complexes with a high molecular weight porcine mannan‐binding protein (pMBP‐28). The α 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 α 2 M/pMBP‐28 complexes was further indicated by crossed immunoelectrophoresis and by use of an anti‐α 2 M affinity column and chelating Sepharose loaded with Zn 2+ . The eluates from these affinity columns showed α 2 M subunits (94 and 180 kDa) and pMBP subunits (28 kDa) in SDS‐PAGE, which reacted with antibodies against α 2 M and pMBP‐28, respectively, in Western blotting. Furthermore, the α 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‐α 2 M reactive material, the other with an M r of 100–150 kDa. The latter peak revealed rhomboid molecules (7 × 15 nm) 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‐α 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 α 2 M.