Activation of Cl, the first component of complement, the generation of Clr-Cls and Cl- inactivator complexes in normal serum by heparin-affinity chromatography

Investigations facilitating heparin-affinity chromatography and immunochemical procedures were utilized to study the interaction and behaviour of heparin with C1, C1 subcomponents and C1&;#x0304; IA in normal serum.Clq bound to heparin-Sepharose independent of divalent cations at 4 and 37° C at physiological pH and salt concentration requiring a high salt concentration for its complete release. MacromolecularC1 bound to heparin-Sepharose in the presence of Ca²⁺ at 37°C, activating Clr and Cls in the process as demonstrated by the appearance of C1&;#x0304;r-C1&;#x0304;s-C1&;#x0304; IA complexes when purified C1&;#x0304; IA was added to the eluted fractions. At 4° C however, proenzyme C1r and C1s were present in the eluted fractions. Proenzyme C1r-C1s and C1&;#x0304;r-C1&;#x0304;s-C1&;#x0304; IA complexes did not bind directly to heparin-Sepharose. Binding of C1r-C1s in the presence of Ca²⁺ was dependent on Clq bound to heparin.High levels of proenzyme C1r-C1s complexes were produced during the 4°C experiments in the presence of Ca²⁺ and were collected in the void volume.C1&;#x0304;r-C1&;#x0304;s-C1&;#x0304; IA complexes generated during the 37°C experiments had no affinity for heparin-Sepharose. C1&;#x0304; IA did not bind directly to heparin. Recirculation of a serum sample collected after Clq was removed by heparinaffinity at 4°C in the presence of EDTA demonstrated that (1) Clr and Cls in the presence of calcium could be reassembled with heparin-bound Clq to form macromolecular Cl. (2) Clr and Cls in non-activated forms did not bind directly to heparin-Sepharose. (3) Clr and Cls contained in the reassembled Cl/heparin complex at 37°C were in active enzymic forms.