Purification and characterization of the C3 convertase of the classical pathway of human complement system by size exclusion high-performance liquid chromatography.
The C3 convertase of the classical pathway of the complement system is a liable complex, C4b,2a, and is activated by limited proteolysis of two components, C4 and C2, by C1s. By utilizing iodine-treated C2 and size exclusion high-performance liquid chromatography (HPLC), we have succeeded in isolating for the first time the classical pathway C3 convertase. Size exclusion HPLC demonstrated that the apparent molecular mass of the C3 convertase was 280K daltons. The C3 convertase decay-dissociates spontaneously into C4b and C2a. The decay-dissociation is a temperature-dependent reaction and the half-lives of the C3 convertase at 24, 30, and 37 degrees C were estimated to be 400, 180, and 60 min, respectively. The decay-dissociation was also dependent on pH and was accelerated by increasing pH. In addition, the decay-dissociation of the C3 convertase was accelerated by C2b. This result suggests that C2b acts as a feedback inhibitor on the activation of the classical pathway of complement system.
D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003166:Complement Activating Enzymes / Q000302:isolation & purification*; D003167:Complement Activation; D003175:Complement C2 / Q000378:metabolism; D050679:Complement C2b; D050577:Complement C3-C5 Convertases / Q000302:isolation & purification*; D003181:Complement C4 / Q000378:metabolism; D015935:Complement C4b; D003171:Complement Pathway, Classical; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008970:Molecular Weight; D013696:Temperature
D003175:Complement C2; D050679:Complement C2b; D003181:Complement C4; D015935:Complement C4b; D003166:Complement Activating Enzymes; D050577:Complement C3-C5 Convertases