![]() ![]() ![]() ![]() The X- ray structure of TIM910 provided no evidence on where or how substrates are bound. We propose that this suppression of dynamics is the underlying principle for the mechanism of action of this antibiotic.In both cases X-ray structures of the apo or antibiotic bound form were available, but not su cient to explain the mechanism of action. The antibiotic cyclomarin completely abolishes dynamics induced by the ligand arginine-phosphate. The N-terminal domain of ClpC1 is the binding site for various new antibiotics against M.tuberculosis. TIM910 binds its substrates in a hy- drophobic pocket on the exterior of the chaperone in a modular fashion, where the number of TIM910 complexes bound depends on the length of the substrate.In the second part I studied the behavior of the N-terminal receptor do- main of the ClpC1 unfoldase from M.tuberculosis in the presence of di erent antibiotics and ligands. Based on nuclear magnetic resonance (NMR), small-angle X-ray scattering (SAXS), analytical ultracentrifugation (AUC) and in vivo mutational experiments I propose a structural model of the chap- erone/membrane protein interaction. Not only is the TIM910 complex in constant exchange between monomeric and hex- americ species, but also the bound substrate samples multiple conformations on a millisecond timescale. In this thesis I present investigations of the structure, dynamics and substrate- interactions of two molecular chaperones, using various biophysical and in vivo methods.In the first part I show that the mitochondrial membrane protein chap- erone TIM910 binds its substrates in a highly dynamic manner. Consequently, structural data on chaperone/substrate complexes are sparse, and the mechanisms of chaperone action are poorly understood. Interactions between the di erent partners in this network and between the substrate and the chaperone are often dynamic processes, which are especially di cult to study using standard structural biology tech- niques. Moreover, molecular chaperones need to be able to interact with each other and with other components of the protein quality control system in a complex network. To this end, molecular chaperones need to be specialized in performing specific tasks, like folding, transport or disaggregation, and versatile in their recognition pattern to engage many di erent client pro- teins. ![]() Ces outils sont actuellement utilisés pour étudier domaines AB de récepteurs nucléaires tels que les RARg, et pour décrire leur modification après phosphorylation.The diverse group of molecular chaperones is dedicated to accompany, fold and protect other proteins until they reach their final conformation and loca- tion inside the cell. Récemment, de nouveaux outils expérimentaux et théoriques ont été développés pour étudier les états perturbés des protéines impliquées dans les mécanismes de signalisation. Souvent, la plasticité moléculaire des protéines est essentielle à leur fonction, comme le démontrent les propriétés d’auto-association de p8, la plus petite des sous-unités de TFIIH ou le rôle des régions avoisinantes des domaines PDZ dans les interactions protéines-peptides. Le groupe est également impliqué dans l’étude de la protéine E6 du papillomavirus humain et de son interaction avec des domaines PDZ de l’hôte. Les questions récemment abordées par notre groupe portent entre autres sur l’étude de plusieurs domaines structurels de facteurs de transcription tels que TFIIH et SAGA. Nous utilisons la RMN, combinée avec d’autres méthodes de biologie structurale, pour étudier les propriétés moléculaires indispensables à la réalisation d’une fonction biologique donnée. La spectroscopie RMN nous donne des aperçus inestimables des caractéristiques structurales et dynamiques des systèmes biomoléculaires. ![]()
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