Significance The complement system discriminates self from nonself targets solely owing to the presence of complement regulators on the host cells, which primarily belong to the regulators of complement activation (RCA) protein family. These regulators control complement activation on the host cell surface by two mechanisms termed “cofactor activity” (CFA) and “decay-accelerating activity.” Here, we have identified the critical structural determinants of an RCA protein responsible for imparting the CFA and show that these determinants bridge MG2 (macroglobulin-2) and CUB (complement C1r-C1s, Uegf, Bmp1) domains of C3b and interact with factor I. As a proof of principle, we show that incorporation of putative membrane cofactor protein (CD46) regions responsible for the bridging and factor I interaction in decay acceleration factor (CD55) results in gain of function. We, thus, define the molecular events that govern CFA.