Purification and physicochemical properties of C1q from guinea-pig serum.
An efficient method is described for the isolation of highly purified, IgG-free and stable guinea-pig serum C1q. The procedure includes the chromatography of EDTA-treated serum (25 mM EDTA) on CM- and DEAE-cellulose followed by gel filtration on ACA 34-Ultrogel whereby ammonium sulfate precipitation was used for concentration. The final product stored in a glycerol containing buffer was purified 700-fold with a yield of approximately 50%. It was judged to be homogeneous by several criteria including SDS-PAGE, analytical ultracentrifugation, gel filtration and immunoprecipitation. The protein has a sedimentation rate of 11.3 S and consists of three distinct polypeptide chains A, B and C with mol. wts of 30,200, 28,200 and 24,000. Amino acid analysis revealed a content of 4.42% hydroxyproline, 1.81% hydroxylysine and 18.7% glycine. In contrast to human serum C1q a very low content of cysteine residues was detected. SDS-PAGE analysis performed in the absence of 2-mercaptoethanol but in the presence of 5-10% SDS revealed clearly that gps-C1q is dissociated in a time-dependent manner into the individual chains.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D003166:Complement Activating Enzymes / Q000302:isolation & purification*; D015922:Complement C1q; D004492:Edetic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D006168:Guinea Pigs; D005779:Immunodiffusion; D014461:Ultracentrifugation
D000596:Amino Acids; D015922:Complement C1q; D004492:Edetic Acid; D003166:Complement Activating Enzymes