Purification and characterization of mouse serum protein with specific binding affinity for C4 (Ss protein)

Abstract

A new component of the complement (C).system, with a specific binding affinity for the activated Ss-protein (C4) has been identified in mouse serum. This protein, named Ss- (or C4)-binding protein (Ss-bp), was purified about 200 times from mouse plasma. Ss-bp is a heat stable (56 degrees C, 60 rain) β-globulin with a sedimentation coefficient in sucrose density ultracentrifugation of 10s. Its concentration in serum of adult male and female mice is 160 and 60 μg/ml, respectively. In EDTA-plasma, Ss and Ss-bp are not associated and can be separated by chromatography in Sephadex G-200. However, in serum Ss-bp binds tightly to Ss. The bonds between these proteins cannot be reversed by chelation of divalent cations. As a consequence of the formation of Ss/Ss-bp complexes, the properties of Ss-bp appear to be quite different in serum of mice with high (Ss-H) or low (Ss-L) levels of Ss-protein. In Ss-H serum, all of Ss- bp is bound to Ss. In Ss-L serum, Ss-bp is mostly free. Because the electrophoretic mobilities of free and complexed Ss-bp are quite different, Ss-bp appears to be polymorphic in serum (but not in EDTA- plasma). The strict dependency of the apparent electrophoretic mobility of Ss-bp on the levels of Ss in serum was demonstrated in a series of congenic mice and among the progeny of a cross between Ss-H and Ss-L strains of mice. Without exception, the slow and fast varieties of Ss-bp were associated with the Ss-L and Ss-H traits. Ss-bp of the slow variety can be transformed into the fast variety by addition of pure human C4, or C4-sufficient guinea pig serum, to Ss-L serum. In both instances Ss-bp formed stable complexes with C4 or a C4- derived peptide. These findings highlight the binding specificity of Ss- bp for the fourth component of the complement system, and in addition they demonstrate a functional homology between the Ss-protein and C4 from two different species.

Human C4-binding protein. Association with immune complexes in vitro and in vivo.

“…Addition of purified C4 to the C4-deficient serum resulted in incorporation of C4-bp and C4. The C4-dependence 2 Fujita, T., and V. Nussenzweig. Manuscript in preparation.…”

Expression and localization of proteins of the complement system in human skin.

“…Both eventuate in the activation ofC5 leading to the formation of the membrane attack complex (2). The system is controlled by serum proteins such as Factor H (3), Factor I (4), C4b binding protein (C4bp) (5), and the membrane proteins C3b receptor (CR 1) (6), C3d receptor (CR2) (7), membrane cofactor protein (MCP) (8), decay accelerating factor (DAF)' (9), and the homologous restriction factors (10). These proteins are widely distributed on the surface of blood and tissue cells.…”

The Oligomerization Domain of C4-Binding Protein (C4bp) Acts as an Adjuvant, and the Fusion Protein Comprised of the 19-Kilodalton Merozoite Surface Protein 1 Fused with the Murine C4bp Domain Protects Mice against Malaria

“…C4bp is an abundant plasma protein first discovered in mice (9). Its natural function is to inhibit the classical and lectin pathways of complement activation (for a review, see reference 3).…”

“…Immunization with the MSP1 19 -murine C4bp fusion protein (referred to below as IMX108) could protect mice against an otherwise lethal parasite challenge, but it also induced antibodies which recognized the host C4bp (9). To eliminate the potential risks associated with such autoimmune antibodies, distantly related homologues of the C4bp domain were tested, and some of these domains proved to be superior in mice to the murine C4bp domain.…”