Isolation and characterization of fibronectin-alpha 1-microglobulin complex in rat plasma.
Molecules containing the 28 kDa immunoregulatory protein alpha 1-microglobulin (alpha 1-m), also known as protein HC, were isolated from rat plasma or serum by immunoaffinity chromatography. Three molecular species were distinguished on the basis of nondenaturing PAGE. Two of these have been described previously: uncomplexed alpha 1-m, and the complex of alpha 1-m with alpha 1-inhibitor-3. The third species was analysed by denaturing PAGE, immunoblotting, proteinase digestion and N-terminal-sequence analyses, and shown to consist of a complex between alpha 1-m and fibronectin. This complex, with a mass of about 560 kDa, was resistant to dissociation in the presence of denaturants, but not in the presence of reducing agents in combination with denaturants, and we conclude that the two components are linked by disulphide bonds. About 60% of the total detectable plasma alpha 1-m exists as high-molecular-mass complexes distributed approximately evenly between fibronectin and alpha 1-inhibitor-3. Immunochemical analyses were used to determine the proportion of the total plasma pools of fibronectin and alpha 1-inhibitor-3 that circulate in complex with alpha 1-m. About 3-7% of the total plasma fibronectin from three different rat strains contained alpha 1-m, whereas 0.3-0.8% of the total plasma alpha 1-inhibitor-3 contained alpha 1-m. Complexes were found at similar levels in plasma and serum, indicating that coagulation is not responsible for complex formation. Moreover, immunochemical analyses of human plasma revealed small amounts of alpha 1-m in complex with fibronectin and alpha 2-macroglobulin (an alpha 1-inhibitor-3 homologue). The existence of a complex between alpha 1-m and fibronectin in rats and humans suggests a mechanism for the incorporation of the immunoregulatory molecule alpha 1-m into the extracellular matrix.
Falkenberg, C; Enghild, JJ; Thøgersen, IB; Salvesen, G; Akerström, B
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