Conformational dynamics of the ABC transporter McjD seen by single-molecule FRET

The EMBO Journal
Florence HusadaThorben Cordes

Abstract

ABC transporters utilize ATP for export processes to provide cellular resistance against toxins, antibiotics, and harmful metabolites in eukaryotes and prokaryotes. Based on static structure snapshots, it is believed that they use an alternating access mechanism, which couples conformational changes to ATP binding (outward-open conformation) and hydrolysis (inward-open) for unidirectional transport driven by ATP Here, we analyzed the conformational states and dynamics of the antibacterial peptide exporter McjD from Escherichia coli using single-molecule Förster resonance energy transfer (smFRET). For the first time, we established smFRET for an ABC exporter in a native-like lipid environment and directly monitor conformational dynamics in both the transmembrane- (TMD) and nucleotide-binding domains (NBD). With this, we unravel the ligand dependences that drive conformational changes in both domains. Furthermore, we observe intrinsic conformational dynamics in the absence of ATP and ligand in the NBDs. ATP binding and hydrolysis on the other hand can be observed via NBD conformational dynamics. We believe that the progress made here in combination with future studies will facilitate full understanding of ABC transport cycles.

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Citations

Apr 21, 2019·Biological Chemistry·Dania Rose-SperlingUte A Hellmich
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Methods Mentioned

BETA
NMR
FRET
confocal
confocal microscopy
Fluorescence

Software Mentioned

LabVIEW
smFRET
PELDOR
ImageQuant TL

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