title: Immunoglobulin M possesses two binding sites for complement subcomponent C1q, and soluble 1:1 and 2:1 complexes are formed in solution at reduced ionic strength.
abstract: Soluble complexes were formed between C1q, a subunit of the first component of human complement, and four different Waldenström IgM proteins at reduced ionic strengths. The equilibria between these complexes and the free proteins were studied in the ultracentrifuge. Complex formation was found to be a very sensitive function of the salt concentration, and at physiological ionic strength complex formation was negligible. The complexes were cross-linked with a water-soluble carbodiimide and separated by sucrose gradient centrifugation. Both 22 S 1:1 and 26 S 2:1 C1q X IgM complexes were formed; stoichiometry was established by cross-linking 125I-C1q with 131I-IgM and determining the ratios of the specific activities of the gradient-purified materials. The association process was studied as a function of protein concentration and was analyzed by Scatchard and Hill plots to yield stoichiometry, association constant, and degree of cooperativity. The results indicated that IgM has two identical and independent binding sites for C1q. The intrinsic association constant was found to vary between 10(6) M-1 at 0.084 M ionic strength to 10(4) M-1 at physiological ionic strength; the slope of the log-log plot gave a value of -6.0. The cross-linked complexes were examined by electron microscopy, and the C1q appeared to be attached to the IgM through the C1q heads, implying that the biologically significant binding sites were involved in this interaction. For the 2:1 complexes, the two C1q appeared to attach to opposite surfaces of the IgM, suggesting the presence of a pseudo-2-fold axis lying in the plane of the IgM disk.
mesh_terms: D001665:Binding Sites; D003166:Complement Activating Enzymes / Q000378:metabolism*; D015922:Complement C1q; D006801:Humans; D007075:Immunoglobulin M / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008433:Mathematics; D008954:Models, Biological; D008958:Models, Molecular; D009994:Osmolar Concentration; D011485:Protein Binding; D011487:Protein Conformation; D012996:Solutions; D008258:Waldenstrom Macroglobulinemia / Q000276:immunology
chemical_list: D007075:Immunoglobulin M; D046911:Macromolecular Substances; D012996:Solutions; D015922:Complement C1q; D003166:Complement Activating Enzymes