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Abstract

The process of translation is mediated by ribosomes, which decode messenger RNA (mRNA) sequences into polypeptide chains. As nascent chains traverse the exit tunnel, their interactions with the tunnel can regulate translation dynamics and co-translational folding. High-resolution ribosome structures have revealed significant geometric variations in the exit tunnel across prokaryotes and eukaryotes. This structural diversity is thought to affect the translation dynamics, particularly of short proteins. To address this, in Chapter 2, I initially applied a simple coarse-grained (CG) molecular dynamics (MD) model to investigate how tunnel geometry in prokaryotes and eukaryotes affects the post-translational escape of proteins ranging from 6 to 56 amino acids (aa) in length. Our simulations revealed a strong protein length-dependent escape behavior that is governed by tunnel geometric constraints. Given the distinct functional roles that tunnel structures play in regulating protein translation across species, I expanded the structural analysis of exit tunnels across a broad range of phylogenetic species in Chapter 3. I refined a developed tunnel-searching pipeline and extracted tunnel geometries from hundreds of cryo-electron microscopy (cryo-EM) ribosome structures across three life domains. I identified six protist species with tunnel geometries that closely resemble those of archaea and bacteria. Using sequence and structure-based comparisons, I revealed that certain modifications in ribosomal proteins and ribosomal RNAs (rRNAs) can explain these similarities, providing new phylogenetic insights into ribosome evolution. Since tunnel structures can vary even within the same domain of life, the simplified tunnel models are insufficient to capture species-specific translation dynamics. Therefore, in Chapter 4, I developed a method to convert tunnel mesh representations into CG bead-based models, enabling efficient simulation of nascent chain dynamics while preserving more accurate tunnel geometry and ribosome surface features. Its utility was further validated by reproducing the known arrest behavior of the VemP peptide. Overall, this thesis offers new insights into the structural heterogeneity of ribosome exit tunnels and its functional implications for translation dynamics of nascent polypeptides; elucidates phylogenetic relationships and ribosome evolution through comparative tunnel structure analysis; and presents a novel simulation framework that can aid in the development of biotechnological strategies targeting protein synthesis.