C9 polymerization has been recognized as the reaction in complement essential for the formation of ultrastructural membrane lesions and cytolysis. In accord with the central role of C9 polymerization during membrane attack, it is proposed to embark on a detailed investigation of the molecular mechanism of C5b-8 mediated C9 polymerization. C5b-8 is the natural C9-polymerizing unit and the elucidation of its mode of action on C9 may enable us to manipulate C9 polymerization in such a way that it may become useful for immunotherapeutic purposes. C5b-8 mediated C9 polymerization will be studied by analyzing the reaction of the first C9 molecule with C5b-8 and determining C9's association partner, conformation, and covalent or noncovalent modification. The primary structure of C9 (and of C7) will be determined by isolating and sequencing C9 (and C7) cDNA clones. The domain structure of C9 and poly C9 will be determined by generating proteolytic fragments of C9, raising antibodies agaist them, and localizing their binding to poly C9. Determination of the primary and domain structure of C9 may enable us to use certain peptide segments of C9 alone or together with domains of C5b-8 as cytolytic effector molecules. C9 polymerization by C5b-8 is under the control of soluble and membrane bound inhibitors. S-protein is the soluble poly C9 inhibitor of plasma. The membrane bound inhibitor(s) on human erythrocyte membranes will be isolated and characterized and its cellular distribution determined. The physiological role of membrane bound poly C9 inhibitors in the protection of """"""""self"""""""" against immunological attack and in the escape of tumor cells from immune destruction will be assessed.
