Modeling of Biomolecular Systems Interactions, Dynamics, and Allostery: Bridging Experiments and Computations - September 10-14, 2014, Istanbul, Turkey

Modeling of Biomolecular Systems Interactions, Dynamics, and Allostery Poster Session II

92-POS Board 45 Going through the Motions: Network Analyses of Molecular Dynamics Simulations Reveal Conserved Intramolecular Communication Pathways within EF-Tu Responsible for Ribosome Dependent GTPase Activation and Nucleotide Exchange. Evan Mercier, Dylan Girodat, Fan Mo, Hans-Joachim Wieden . University of Lethbridge, Lethbridge, Canada. During the elongation cycle of bacterial protein synthesis elongation factor (EF) Tu delivers aminoacyl(aa)-tRNAs to the ribosome in a GTP-dependent manner. EF-Tu functions as a checkpoint, correct codon recognition is required to send an activating signal from the decoding centre on the 30S ribosomal subunit to the G domain of EF-Tu bound to the 50S ribosomal subunit. Following dissociation, EF-Tu•GDP is recycled into its GTP-bound form through the interaction with its nucleotide exchange factor EF-Ts. Both functions require the transmission of signals to facilitate either GTP hydrolysis or nucleotide release. Little is known about the dynamic features governing signal transmission within a highly conserved protein such as EF-Tu and its particular evolutionary constraints. Here, molecular dynamics simulations of EF-Tu and variants with single amino acid substitutions were performed and subsequently interrogated using network analysis methods. Using these networks we identify communication pathways essential for efficient GTPase activation on the ribosome as well as EF-Ts-catalyzed nucleotide exchange. We find that single amino acid substitutions in EF-Tu can change the network organization dramatically and result in significantly reduced communication between domains across a set of universally conserved interdomain bridges. To validate our findings we have performed Michaelis-Menten analyses of ribosome-stimulated GTP hydrolysis in EF-Tu. Consistent with our network analysis, these results reveal that amino acid substitutions in domain II reduce the stimulatory effect of the 70S ribosome five-fold, while not affecting 50S-dependent stimulation. Variants of EF-Tu that disrupt the interdomain bridges reduce the stimulatory effect of the 70S ribosome to a similar extent. Analyses of the EF-Tu•EF-Ts complex revealed a different set of communication pathways promoting nucleotide exchange in EF-Tu which were validated using rapid-kinetics techniques.

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