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@ARTICLE{Viennet:862750,
author = {Viennet, Thibault and Bungert-Plümke, Stefanie and Elter,
Shantha and Viegas, Aldino and Fahlke, Christoph and
Etzkorn, Manuel},
title = {{R}econstitution and {NMR} {C}haracterization of the
{I}on-{C}hannel {A}ccessory {S}ubunit {B}arttin in
{D}etergents and {L}ipid-{B}ilayer {N}anodiscs},
journal = {Frontiers in molecular biosciences},
volume = {6},
issn = {2296-889X},
address = {Lausanne},
publisher = {Frontiers},
reportid = {FZJ-2019-02992},
pages = {13},
year = {2019},
abstract = {Barttin is an accessory subunit of ClC-K chloride channels
expressed in the kidney and the inner ear. Main functions of
ClC-K/barttin channels are the generation of the
cortico-medullary osmotic gradients in the kidney and the
endocochlear potential in the inner ear. Mutations in the
gene encoding barttin, BSND, result in impaired urinary
concentration and sensory deafness. Barttin is predicted to
be a two helical integral membrane protein that directly
interacts with its ion channel in the membrane bilayer where
it stabilizes the channel complex, promotes its
incorporation into the surface membrane and leads to channel
activation. It therefore is an attractive target to address
fundamental questions of intermolecular communication within
the membrane. However, so far inherent challenges in protein
expression and stabilization prevented comprehensive in
vitro studies and structural characterization. Here we
demonstrate that cell-free expression enables production of
sufficient quantities of an isotope-labeled barttin variant
(I72X Barttin, capable to promote surface membrane insertion
and channel activation) for NMR-based structural studies.
Additionally, we established purification protocols as well
as reconstitution strategies in detergent micelles and
phospholipid bilayer nanodiscs. Stability, folding, and NMR
data quality are reported as well as a suitable assignment
strategy, paving the way to its structural
characterization.},
cin = {ICS-6 / ICS-4},
ddc = {570},
cid = {I:(DE-Juel1)ICS-6-20110106 / I:(DE-Juel1)ICS-4-20110106},
pnm = {551 - Functional Macromolecules and Complexes (POF3-551) /
553 - Physical Basis of Diseases (POF3-553)},
pid = {G:(DE-HGF)POF3-551 / G:(DE-HGF)POF3-553},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:30931313},
UT = {WOS:000463667700001},
doi = {10.3389/fmolb.2019.00013},
url = {https://juser.fz-juelich.de/record/862750},
}