MEX-3C, a novel RNA binding E3 ubiquitin ligases, contains two N-terminal heterogeneous nuclear ribonucleoprotein K homology (KH) domains and C-terminal Ring finger domain. Recent evidence has suggested that human MEX-3C has a strong bondage with carcinogenesis and the MEX-3C-mediated ubiquitination of RIG-I is essential for the antiviral innate immune response. Moreover, the Ring finger domain of MEX-3C could regulate the degradation of HLA-A2 (an MHC-I allotype) mRNA with a novel mechanism. However, the structural basis for the ubiquitination catalyzed by hMEX-3C Ring finger domain remains evasive. In this study, we solved the crystal structure of dimeric Ring finger domain of hMEX-3C and compared it with the complex structure of MDM2/MDMX-UbcH5b-Ub. Our ubiquitination assay demonstrated that the Ring finger domain of hMEX-3C acts as a ubiquitin E3 ligase in vitro, cooperating with specific E2 to mediate ubiquitination. Then, we identified several key residues in Ring finger domain of hMEX-3C possibly involved in the interaction with E2-Ub conjugate and analyzed the E3 ligase activities of wild type and mutants at key sites. Additionally, zinc chelation experiments indicated that the intact structural stability is essential for the self-ubiquitination activity of the Ring finger domain of hMEX-3C. Taken together, our studies provided new insight into the mechanism of the Ring finger domain of hMEX-3C that may play an important role in eliciting antiviral immune responses and therapeutic interventions.
Keywords: E3 ubiquitin ligase; Ring finger domain; hMEX-3C; ubiquitination.
© 2018 The Authors. Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.