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001024924 0247_ $$2datacite_doi$$a10.34734/FZJ-2024-02574
001024924 0247_ $$2URN$$aurn:nbn:de:0001-20240516082247276-4907644-5
001024924 020__ $$a978-3-95806-754-7
001024924 037__ $$aFZJ-2024-02574
001024924 1001_ $$0P:(DE-Juel1)180391$$aBustorff, Nuno$$b0$$eCorresponding author$$ufzj
001024924 245__ $$aFolding and structural studies of saccharomyces cerevisiae Phosphoglycerate Kinase$$f- 2024-04-30
001024924 260__ $$aJülich$$bForschungszentrum Jülich GmbH Zentralbibliothek, Verlag$$c2024
001024924 300__ $$axxvi, 126
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001024924 3367_ $$0PUB:(DE-HGF)11$$2PUB:(DE-HGF)$$aDissertation / PhD Thesis$$bphd$$mphd$$s1712897890_11837
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001024924 4900_ $$aSchriften des Forschungszentrums Jülich Reihe Schlüsseltechnologien / Key Technologies$$v282
001024924 502__ $$aDissertation, RWTH Aachen University, 2024$$bDissertation$$cRWTH Aachen University$$d2024
001024924 520__ $$aProteins are synthesized within cells by ribosomes, and their functionality depend on the correct three-dimensional structure obtained through the process of folding. While the classical understanding of protein folding primarily focused on postsynthesisfolding, recent research has shifted its emphasis toward unraveling the intricacies of folding during synthesis. Of particular interest in this pursuit are multi-domain proteins, which constitute over 70% of proteins in cells. Using classical single-molecule fluorescence resonance transfer efficiency (sm-FRET) studies, the unfolding/refolding transitions of a two-domain yeast phosphoglycerate kinase (yPGK) was explored as a model for multiple domain proteins. To enhance our understanding of a transition within a single-domain of full-length yPGK, I assessed two FRET pair variants within the N-terminal domain. Together with previous data we compared in total six different variants, for which we observed three distinct transitions in both domains: the first segment (positions 1–88 and 202–256, respectively) showed no transition (i.e. no change in the distance), while the second segment (positions 34–135 and 256–290, respectively) underwent a conventional two-state transition. Intriguingly, labeling the complete Rossmann motif (positions 1–135 and 202–290, respectively) revealed a compact intermediate state during GuHCl-induced unfolding in transitory conditions. The detailed understanding of N-terminal domain transitions was essential for comparative analyses with shorter-length proteins synthesized for co-translational folding studies. In order to investigate the ribosomal folding process of yPGK, I employed cryo-electron microscopy (cryo-EM). Four distinct ribosome nascent chain complex (RNC) structures were solved, each representing a nascent polypeptide of a varying length. Most structures depicted nascent chain density outside the ribosomal tunnel, and we observed for the first time the structure of a full-length protein nascent chain attached to the ribosome. In summary, the presented research advances our knowledge of yPGK folding transitions and offers novel routes for studying co-translational folding processes within RNC complexes using sm-FRET and cryo-EM.
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001024924 536__ $$0G:(DE-HGF)POF4-5241$$a5241 - Molecular Information Processing in Cellular Systems (POF4-524)$$cPOF4-524$$fPOF IV$$x1
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001024924 9141_ $$y2024
001024924 9101_ $$0I:(DE-588b)5008462-8$$6P:(DE-Juel1)180391$$aForschungszentrum Jülich$$b0$$kFZJ
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