Isolation and biochemical characterization of alpha-2-opsonic glycoprotein from rat serum.

The importance of a circulating a-2-globulin as a determinant of hepatic Kupffer cell phagocytosis of nonbacterial particulate matter has been well documented. This protein which manifests opsonic activity exerts a humoral control on hepatic reticuloendothelial phagocytic activity and deficiency of its serum level exists with advanced malignant diseases or acutely following traumatic injury. The present study was designed to isolate this cu-2-opsonic protein from rat serum by using a combination of gel filtration in 6 M urea on cross-linked Sepharose 6B with prior sequential steps of ammonium sulfate fractionation, high voltage preparative free flow electrophoresis, and conventional Sepharose 4B gel filtration. Immunochemical analysis utilizing nonabsorbed antiserum to the rat opsonic protein as well as polyacrylamide gel electrophoresis was used to evaluate the level of purification, which was in excess of 99%. The purified protein was selectively immunoreactive with monospecific antiserum and its serum concentration was quantified by electroimmunoassay. The protein consists of two subunits held together by disulfide bonds and the minimum molecular weight as determined by sodium dodecyl sulfatepolyacrylamide gel electrophoresis of each subunit was 2.29 x lo”. Amino acid analysis revealed no detectable methionine and a high concentration of hydrophilic amino acid residues. Amino sugar analysis confirmed its classification as a glycoprotein with approximately 1.8% glucosamine and no galactosamine. The purification and biochemical analysis of this cY-2-glycoprotein may provide insight as to its mechanism of action with respect to reticuloendothelial function.