We have modified our published procedure for the rapid isolation of the second component of human complement to obtain increased yield and specific functional activity. Also isolated by this new method is a protein which has many physicochemical properties in common with C2. See project #Z01 AI 00271-06 LCI for a detailed description of the isolation and partial characterization of this new plasma protein termed gp120 kDa. The capacity of this protein to specifically bind to iC4/C4b, the numerous similarities to C2 and its relatively high plasma concentration (250-300 ug/ml) strongly suggested that this sialoglycoprotein may participate in the classical complement pathway. Thus, initial attempts to delineate a functional role for gp120 kDa were directed to the early events of complement activation in this pathway. We used complement coated sheep erythrocytes with limited functional sites EAC4b or EAC1, 4b as well as one or more other limited complement components to study the effect of gp120 kDa at different points in the formation of the classical pathway C5 convertase. By several distinct functional tests we were able to show dramatic inhibition (>90%) by gp120 kDa of classical pathway convertase formation. Inhibition was shown for C1, C2, and C3 site formation and up to 30% acceleration of C2 decay was observed. The exact point(s) where gp120 kDa exerts its influence has to be determined possibly by competition and binding experiments using radiolabeled proteins. These effects were obtained at input concentrations between 15 and 255 ug/ml of gp120 kDa. To assess the ability of gp120 kDa to functionally influence later steps in the complement sequence we also studied its affect on reactive lysis. Again this glycoprotein significantly inhibited complement mediated lysis both during and following EAC5b,6,7 site formation. Since we have shown the direct binding of gp120 kDa to surface bound iC4/C4b, we did expect an influence on the convertase formation. However, the ability of gp120 kDa to inhibit reactive lysis was unexpected and further work is needed elucidate the mode of action on the terminal complement components.
