Formation and Functional Significance of a Molecular Complex Derived From the Second and the Fourth Component of Human Complement

Abstract

Studies on the mechanism of immune cytolysis have been greatly facifitated by the use of highly purified complement components. Paradoxically much can be learned about immune cytolysis by studying the reactions of complement components in solutions which are entirely devoid of cells. The experiments described in the present paper were performed to explore the molecular interactions between the first four components of human complement in cell-and antibody-free solution. Use was made of highly purified preparations of the esterase moiety of the first component (C'1), and of the second (C'2), third (C'3), and fourth (C'4) component. It was found that C'2 and C'4 tend to interact in free solution to form a reversible protein-protein complex which is converted into a stable complex by the action of C'I esterase. Formation of the esterase-induced complex results in the generation of an enzyme-like activity which causes the conversion of CP3 to its hemolytically inactive product, Cr3i (1). Evidence will be presented indicating that the complex is formed not only in cell-free solution, but also on the surface of cells and that it fulfills an essential function in cell membrane damage by complement. Materials and MethodsPurified Human Complement Fa~tors.--The esterase moiety of the first component of complement (2) was isolated and kindly provided by Dr. Irwin Lepow, Cleveland, Ohio.The second component was purified from the hydrazine-trested pseudoglobniin fraction of human serum by a three step procedure which will be described in detail elsewhere. 1 Briefly, the first step consists of chromatography on citrboxymethyl-ceUniose using phosphate buffer, pH 6.0, ionic strength 0.05 containing 0.0025 ~ EDTA. The protein is eluted by an increasing * This is publication No. 190 from

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