BLaTM 2.0, a Genetic Tool Revealing Preferred Antiparallel Interaction of Transmembrane Helix 4 of the Dual-Topology Protein EmrE

Julius, Ayse; Laur, Lisa; Schanzenbach, Christoph; Langosch, Dieter

doi:10.1016/j.jmb.2017.04.003

Cited by 6 publications

(13 citation statements)

References 35 publications

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“…GldL-TMH1 and GldM-TMH specifically formed homodimers but no interaction was detected between GldL-TMH1 and GldM TMH ( Fig 2C ). To test the interaction with GldL-TMH2, which exhibits an out-to-in topology, we used the BLA assay [ 56 , 57 ]. GldL-TMHs and GldM-TMH were fused to either the N- or carboxyl-terminal domain of β-lactamase (Bla).…”

Section: Resultsmentioning

confidence: 99%

Dynamic proton-dependent motors power type IX secretion and gliding motility in Flavobacterium

et al. 2022

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Motile bacteria usually rely on external apparatus like flagella for swimming or pili for twitching. By contrast, gliding bacteria do not rely on obvious surface appendages to move on solid surfaces. Flavobacterium johnsoniae and other bacteria in the Bacteroidetes phylum use adhesins whose movement on the cell surface supports motility. In F. johnsoniae, secretion and helicoidal motion of the main adhesin SprB are intimately linked and depend on the type IX secretion system (T9SS). Both processes necessitate the proton motive force (PMF), which is thought to fuel a molecular motor that comprises the GldL and GldM cytoplasmic membrane proteins. Here, we show that F. johnsoniae gliding motility is powered by the pH gradient component of the PMF. We further delineate the interaction network between the GldLM transmembrane helices (TMHs) and show that conserved glutamate residues in GldL TMH are essential for gliding motility, although having distinct roles in SprB secretion and motion. We then demonstrate that the PMF and GldL trigger conformational changes in the GldM periplasmic domain. We finally show that multiple GldLM complexes are distributed in the membrane, suggesting that a network of motors may be present to move SprB along a helical path on the cell surface. Altogether, our results provide evidence that GldL and GldM assemble dynamic membrane channels that use the proton gradient to power both T9SS-dependent secretion of SprB and its motion at the cell surface.

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Section: Resultsmentioning

confidence: 99%

Dynamic proton-dependent motors power type IX secretion and gliding motility in Flavobacterium

et al. 2022

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“…Structural studies of EmrE using NMR spectroscopy ( 8 , 9 ), cryo-electron microscopy (cryo-EM) ( 5 ), and X-ray crystallography ( 10 ) have established two conformationally distinguishable monomeric states that are simultaneously populated within each dimer ( 11 ). Förster resonance energy transfer (FRET) experiments have conclusively demonstrated the antiparallel orientation of the monomers with respect to one another ( 8 ), which may be due to the more favorable helix–helix interactions in antiparallel orientations relative to parallel ones ( 12 ). Based on this evidence, proposed transport cycles for EmrE proceed by an alternating access mechanism ( 13 , 14 ), whereby the monomers swap conformations between these two states, as in other transporters ( Fig.…”

mentioning

confidence: 99%

Electrostatic lock in the transport cycle of the multidrug resistance transporter EmrE

Vermaas

Rempe

Tajkhorshid

2018

Proc. Natl. Acad. Sci. U.S.A.

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SignificanceEmrE is a small membrane transporter found in Escherichia coli that exports drug-like molecules from the cell, contributing to antibiotic resistance. In EmrE, as well as in the wider small-multidrug resistance transporter family, a specific anionic amino acid (E14) has been implicated in governing the conformational changes that export drugs. However, due to sparse structural information, the exact interactions remain unidentified. Through interactive molecular dynamics to incorporate existing cryo-electron microscopy data, we create a fully refined atomic model of EmrE. We then embed this model in a lipid bilayer and evaluate the interactions within EmrE under different loading states. We find that E14 makes specific hydrogen bonds to neighboring residues, coupling observed experimental phenomena to interactions at the atomic scale.

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“…2C). To test the interaction with GldL-TMH2, which exhibits an out-to-in topology, we used the BLA assay (Julius, Laur et al 2017, Schanzenbach, Schmidt et al 2017.…”

Section: Gldl and Gldm Constitute The Molecular Motor That Couples Pmf To Gldm Conformational Changesmentioning

confidence: 99%

Dynamic proton-dependent motors power Type IX secretion and gliding adhesin movement inFlavobacterium

Vincent

Hervada

Sebban‐Kreuzer

et al. 2021

Preprint

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Motile bacteria usually rely on external apparatus like flagella for swimming or pili for twitching. By contrast, gliding bacteria do not rely on obvious surface appendages to move on solid surfaces. Flavobacterium johnsoniae and other bacteria in the Bacteroidetes phylum use adhesins whose movement on the cell surface supports motility. In F. johnsoniae, secretion and helicoidal motion of the main adhesin SprB are intimately linked and depend on the type IX secretion system (T9SS). Both processes necessitate the proton motive force (PMF), which is thought to fuel a molecular motor that comprises the GldL and GldM cytoplasmic membrane proteins. Here we show that F. johnsoniae gliding motility is powered by the pH gradient component of the PMF. We further delineate the interaction network between the GldLM transmembrane helices (TMH) and show that conserved glutamate residues in GldL TMH are essential for gliding motility, although having distinct roles in SprB secretion and motion. We then demonstrate that the PMF and GldL trigger conformational changes in the GldM periplasmic domain. We finally show that multiple GldLM complexes are distributed in the membrane suggesting that a network of motors may be present to move SprB along a helical path on the cell surface. Altogether, our results provide evidence that GldL and GldM assemble dynamic membrane channels that use the proton gradient to power both T9SS-dependent secretion of SprB and its motion at the cell surface.

show abstract

BLaTM 2.0, a Genetic Tool Revealing Preferred Antiparallel Interaction of Transmembrane Helix 4 of the Dual-Topology Protein EmrE

Cited by 6 publications

References 35 publications

Dynamic proton-dependent motors power type IX secretion and gliding motility in Flavobacterium

Dynamic proton-dependent motors power type IX secretion and gliding motility in Flavobacterium

Electrostatic lock in the transport cycle of the multidrug resistance transporter EmrE

Dynamic proton-dependent motors power Type IX secretion and gliding adhesin movement inFlavobacterium

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