Structural basis of Rho GTPase recognition by C3 exoenzyme

C3 exoenzyme is a mono-ADPribosyltransferase (ART) that catalyzes transfer of an ADP-ribose moiety from NAD to Rho GTPases. C3 has long been used to study the diverse regulatory functions of Rho GTPases. How C3 recognizes its substrate and ADP-ribosylation proceeds are still poorly understood. Crystal structures of C3-RhoA complex reveal that C3 recognizes RhoA via switch I, switch II and interswitch regions. In C3-RhoA(GTP) and C3-RhoA(GDP), switch I and II adopt the GDP and GTP conformations, respectively, which explains why C3 can ADPribosylate both nucleotide forms. Based on structural information, we successfully changed Cdc42 to active substrate with combined mutations in the C3-Rho GTPase interface. Moreover, the structure reflects the close relationship among Gln183 in the QXEmotif (C3), a modified Asn41 residue (RhoA) and NC1 of NAD(H), which suggests C3 is the prototype ART. The structures show directly for the first time that the ARTT-loop is the key to target protein recognition and also serve to bridge the gaps among independent studies of