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Tassis, Konstantinos; Vietrov, Ruslan; Koning, Matthijs de; Boer, Marijn de; Gouridis, Giorgos and Cordes, Thorben ORCID logoORCID: https://orcid.org/0000-0002-8598-5499 (2021): Single-molecule studies of conformational states and dynamics in the ABC importer OpuA. In: FEBS letters, Vol. 595, No. 6: pp. 717-734 [PDF, 1MB]


The current model of active transport via ABC importers is mostly based on structural, biochemical and genetic data. We here establish single-molecule Förster resonance energy transfer (smFRET) assays to monitor the conformational states and heterogeneity of the osmoregulatory type I ABC importer OpuA from Lactococcus~lactis. We present data probing both intradomain distances that elucidate conformational changes within the substrate-binding domain (SBD) OpuAC, and interdomain distances between SBDs or transmembrane domains. Using this methodology, we studied ligand-binding mechanisms, as well as ATP and glycine betaine dependences of conformational changes. Our work expands the scope of smFRET investigations towards a class of so far unstudied ABC importers, and paves the way for a full understanding of their transport cycle in the future.

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