Revealing an outward-facing open conformational state in a CLC Cl(-)/H(+) exchange transporter

ELife
Chandra M KhantwalMerritt C Maduke

Abstract

CLC secondary active transporters exchange Cl(-) for H(+). Crystal structures have suggested that the conformational change from occluded to outward-facing states is unusually simple, involving only the rotation of a conserved glutamate (Gluex) upon its protonation. Using (19)F NMR, we show that as [H(+)] is increased to protonate Gluex and enrich the outward-facing state, a residue ~20 Å away from Gluex, near the subunit interface, moves from buried to solvent-exposed. Consistent with functional relevance of this motion, constriction via inter-subunit cross-linking reduces transport. Molecular dynamics simulations indicate that the cross-link dampens extracellular gate-opening motions. In support of this model, mutations that decrease steric contact between Helix N (part of the extracellular gate) and Helix P (at the subunit interface) remove the inhibitory effect of the cross-link. Together, these results demonstrate the formation of a previously uncharacterized 'outward-facing open' state, and highlight the relevance of global structural changes in CLC function.

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Citations

Feb 18, 2016·Journal of the American Chemical Society·Tao JiangEmad Tajkhorshid
Oct 27, 2016·Journal of the American Chemical Society·Sangyun LeeGregory A Voth
Dec 22, 2016·Nature·Eunyong ParkRoderick MacKinnon
May 31, 2018·Physiological Reviews·Thomas J Jentsch, Michael Pusch
May 29, 2018·ELife·Eunyong Park, Roderick MacKinnon
Jun 12, 2018·ELife·Erika A RiedererFrancis I Valiyaveetil
Oct 22, 2019·Journal of Computational Chemistry·Zhi WangGregory A Voth
Apr 26, 2019·PLoS Biology·Kaituo WangPontus Gourdon
Mar 2, 2017·The Journal of General Physiology·Malvin VienAlessio Accardi
Jun 27, 2018·Nature Structural & Molecular Biology·Nicholas B LastChristopher Miller
Aug 15, 2019·Proceedings of the National Academy of Sciences of the United States of America·Kunwoong ParkHyun-Ho Lim
Feb 19, 2021·The Journal of General Physiology·Chris Miller
Sep 7, 2017·Biophysical Journal·Cat Chenal, M R Gunner
Feb 23, 2021·Journal of Molecular Biology·Kacey MerschJanice L Robertson
May 6, 2021·The Journal of Physical Chemistry Letters·Maria Gabriella ChiarielloPaolo Carloni
Jun 18, 2021·Nature·George R HeathSimon Scheuring
Jan 16, 2019·Journal of Chemical Theory and Computation·Adam W DusterHai Lin
Jan 16, 2018·Journal of the American Chemical Society·Heather B MayesJessica M J Swanson

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Methods Mentioned

BETA
cross-linking studies
NMR
X-ray
size exclusion chromatography
gel filtration
isothermal titration calorimetry
PCA
size
gel
ion-exchange chromatography

Software Mentioned

refmac
VMD
XDS
CHARMM
Origin
Topspin
NIH ImageJ
HOLE
CHARMM36
MOLREP

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