Investigating the mechanisms of protein synthesis using multi-resolution structural data

摘要

The ribosome is a complex, dynamic molecular machine responsible for protein synthesis in all cells according to the genetic information. Recent breakthroughs in ribosome crystallography culminated with the 2009 Nobel Prize in Chemistry. Concomitantly, advances in cryo-electron microscopy (cryo-EM) enabled the determination of images of the ribosome trapped in functional states at ever increasing resolution. In order to study different aspects of ribosome function at the atomic level, we developed the molecular dynamics flexible fitting (MDFF) method that combines X-ray and cryo-EM data, furnishing atomic models of the ribosome corresponding to functional intermediates. The MDFF-derived atomic models, combined with molecular dynamics simulations and other computational techniques, allowed us to address different research questions presented in this thesis. First, we found how ribosome-induced changes in the structure of elongation factor Tu leads to its GTPase activation, a crucial step in the decoding of genetic information. Next, we investigated structural and regulatory aspects of ribosomes in complex with a protein-conducting channel, which transports certain nascent proteins across or into membranes. Another area of investigation was the recognition of a regulatory nascent chain by the ribosome, as well as the mechanism by which it leads to translational stalling. Finally, we studied intermediate states of translocation of messenger and transfer RNAs through the ribosome, reconciling data from cryo-EM and single-molecule experiments.