Dynamics and Energetics of Elongation Factor SelB in the Ternary Complex and the Ribosome

Abstract

SelB is an elongation factor specialized to deliver the selenocysteine (Sec) tRNA to the ribosome by recoding the UGA stop codon on the mRNA. Initially the tRNA is in complex with selB and GTP forming the ternary complex (TC). High-resolution cryo-EM structures of intermediates of the Sec incorporation pathway uncover large-scale conformational changes of the ribosome and the TC. To complement the structural information with energetics and rapid dynamics, we performed extensive all-atom molecular dynamics simulations of the ribosome with bound TC as well as of the free TC in solution. The simulations of the free TC were started after extracting the TC from the ribosome-bound cryo-EM structures. The TC was found to rapidly interconvert between the different conformations allowing us to construct the free-energy landscape of the involved motions. This free-energy landscape indicates that the intrinsic large-scale conformational changes of the tRNA and SelB during the delivery to the ribosome are not rate-limiting to the process. In simulations of the free TC started from the GTPase-activated ribosome-bound conformation, the TC rapidly transitions into an inactivated conformation, showing that the GTPase-activated state is strongly stabilized by the ribosome. The simulations of the full ribosome with bound TC in the intermediate states allow us to identify the motions that are rate-limiting to the process of tRNA delivery and to identify the molecular mechanism of the domain closure of small ribosomal subunit upon tRNA decoding.