Structural elucidation of ribosomal protection mediated by MsrE

Abstract

The rapid development of antibiotics resistance crisis and the lack of structures which implicate ribosomal protection by the ubiquitous ABC-F proteins warrants research. MsrE is a horizontal gene transferable member of ATP Binding Cassette protein-F (ABC-F) family which confers cross-species antibiotics resistance to macrolides and streptogramin B. Structural elucidation of MsrE would give us important first structural insights towards building a model for ABC-F mediated ribosome protection. Biochemical, mutational, microbiological and structural investigations were performed to understand the molecular mechanism by which MsrE mediates macrolide resistance. The Cryo-EM structure of ribosome-bound MsrE was successfully determined at 3.6 Å, which enabled its structural modelling despite the lack of MsrE crystal structure. Interestingly, it is revealed that MsrE binds AMP-PNP, which dimerizes its ABC domains into a “closed” conformation. MsrE in “closed” conformation binds from the E-site of ribosome. From there, MsrE domain linkers and extended loop are inserted towards the Peptidyl Transferase Center of the ribosome. Comparison with azithromycin-bound ribosome reveals conformational changes to azithromycin binding site and structural clash between the extended loop of MsrE and azithromycin. ATP hydrolysis is necessary for in vivo resistance, but not for in vitro displacement of azithromycin implying its significance in MsrE turnover from ribosome binding. Based on experimental data and literature review, a model is proposed for antibiotic resistance element (ARE) ABC-F mediated ribosome protection. Preliminary work on a MsrE inhibitor towards Structure-Based Drug Design (SBDD) is also reported.