The dynamic cycle of bacterial translation initiation factor IF3

Initiation factor IF3 is an essential protein that enhances the fidelity and speed of bacterial mRNA translation initiation. Here, we describe the dynamic interplay between IF3 domains and their alternative binding sites using pre-steady state kinetics combined with molecular modelling of available structures of initiation complexes. Our results show that IF3 accommodates its domains at velocities ranging over two orders of magnitude, responding to the binding of each 30S ligand. IF1 and IF2 promote IF3 compaction and the movement of the C-terminal domain (IF3C) towards the P site. Concomitantly, the N-terminal domain (IF3N) creates a pocket ready to accept the initiator tRNA. Selection of the initiator tRNA is accompanied by a transient accommodation of IF3N towards the 30S platform. Decoding of the mRNA start codon displaces IF3C away from the P site and rate limits translation initiation. 70S initiation complex formation brings IF3 domains in close proximity to each other prior to dissociation and recycling of the factor for a new round of translation initiation. Altogether, our results describe the kinetic spectrum of IF3 movements and highlight functional transitions of the factor that ensure accurate mRNA translation initiation. InnovatePeru [382-PNICP-PIBA-2014 and 297INNOVATEPERU-EC-2016 to P.M.]; Fondo Nacional de Desarrollo Cientifico, Tecnologico y de Innovacion Tecnologica [154-2017-Fondecyt and 0362019-Fondecyt-BM-INC.INV to P.M.]; FIRB Futuro in Ricerca [RBFR130VS5 001 to A.F.]; Italian Ministero dell'Istruzione, dell'Universita e della Ricerca (to A.F.); Part of the work on structural dynamics of the ribosome was supported by Russian Science Foundation [17-1401416 to A.L.K.]. Funding for open access: Universidad Peruana de Ciencias Aplicadas (Exp-03).