The reaction of [14C]methylamine with native human C3 led to the stoichiometric incorporation of methylamine, loss of hemolytic activity, and the concomitant exposure of a sulfhydryl group that could be labeled with [14C]iodoacetamide. Both labeled sites were located in the C3d portion of the alpha-chain, which is known to contain the metastable binding of C3b. The methylamine-modified C3 [C3(CH3NH2)] was shown to exhibit many of the functional properties of C3b, although the C3a portion of the molecule remained covalently attached. C3(CH3NH2) bound Factor B and beta 1H, and could be cleaved by C3b inactivator in the presence of beta 1H. C3(CH3NH2) added to human serum caused activation of the alternative pathway and consumption of C3. In presence of Factors B and D and Mg++, C3(CH2NH2) formed a C3 convertase. The convertase-forming material could be removed from solution by anti-C3a Sepharose and the preformed convertase was completely inhibited by purified antibody to C3a. This antibody did not affect the function of the C3 convertase that contained C3b. Similar functional properties were exhibited by C3 exposed for short periods of time to relatively low concentrations of chaotropic reagents, such as KSCN or guanidine. These results suggest that the initial C3 convertase of the alternative pathway may be formed from native C3, without proteolysis, by the attack of a variety of nucleophiles including water. The C3 convertase formed from this altered C3 then generates by proteolytic cleavage the initial metastable C3b that is capable of attaching to receptive surfaces. Conversion of C3 to C3b exposes one sulfhydryl residue as does modification of C3 with methylamine. When the C3d portion of C3b bound to zymosan particles via the metastable binding site was treated with radiolabeled methylamine, the fragment was released from the particles in radiolabeled form. These findings are consistent with the concept that native C3 contains an active carbonyl group, probably in the form of a thioester, which can either react with water to form functionally C3b-l;ike C3 or, upon enzymatic conversion of C3 to C3b, allows C3b to form an ester bond with hydroxyl groups on the target surface.