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dc.contributor.author
Mireku, Samantha A.
dc.contributor.author
Ruetz, Markus
dc.contributor.author
Zhou, Tianyu
dc.contributor.author
Korkhov, Vladimir M.
dc.contributor.author
Kräutler, Bernhard J.
dc.contributor.author
Locher, Kaspar P.
dc.date.accessioned
2018-10-12T14:56:28Z
dc.date.available
2017-06-12T19:43:41Z
dc.date.available
2018-10-12T14:56:28Z
dc.date.issued
2017-01-27
dc.identifier.issn
2045-2322
dc.identifier.other
10.1038/srep41575
en_US
dc.identifier.uri
http://hdl.handle.net/20.500.11850/128405
dc.identifier.doi
10.3929/ethz-b-000128405
dc.description.abstract
BtuCD-F is an ABC transporter that mediates cobalamin uptake into Escherichia coli. Early in vivo data suggested that BtuCD-F might also be involved in the uptake of cobinamide, a cobalamin precursor. However, neither was it demonstrated that BtuCD-F indeed transports cobinamide, nor was the structural basis of its recognition known. We synthesized radiolabeled cyano-cobinamide and demonstrated BtuCD-catalyzed in vitro transport, which was ATP- and BtuF-dependent. The crystal structure of cobinamide-bound BtuF revealed a conformational change of a tryptophan residue (W66) in the substrate binding cleft compared to the structure of cobalamin-bound BtuF. High-affinity binding of cobinamide was dependent on W66, because mutation to most other amino acids substantially reduced binding. The structures of three BtuF W66 mutants revealed that tight packing against bound cobinamide was only provided by tryptophan and phenylalanine, in line with the observed binding affinities. In vitro transport rates of cobinamide and cobalamin were not influenced by the substitutions of BtuF W66 under the experimental conditions, indicating that W66 has no critical role in the transport reaction. Our data present the molecular basis of the cobinamide versus cobalamin specificity of BtuCD-F and provide tools for in vitro cobinamide transport and binding assays.
en_US
dc.format
application/pdf
en_US
dc.language.iso
en
en_US
dc.publisher
Nature
dc.rights.uri
http://creativecommons.org/licenses/by/4.0/
dc.subject
X-ray crystallography
en_US
dc.subject
Transporters
en_US
dc.subject
Protein transport
en_US
dc.subject
Membrane proteins
en_US
dc.subject
Biocatalysis
en_US
dc.title
Conformational Change of a Tryptophan Residue in BtuF Facilitates Binding and Transport of Cobinamide by the Vitamin B12 Transporter BtuCD-F
en_US
dc.type
Journal Article
dc.rights.license
Creative Commons Attribution 4.0 International
ethz.journal.title
Scientific Reports
ethz.journal.volume
7
en_US
ethz.journal.abbreviated
Sci Rep
ethz.pages.start
41575
en_US
ethz.size
11 p.
en_US
ethz.version.deposit
publishedVersion
en_US
ethz.grant
Structural and mechanistic studies of components of bacterial protein N-glycosylation pathway and of vitamin B12 transport
en_US
ethz.identifier.wos
ethz.identifier.scopus
ethz.identifier.nebis
006751867
ethz.publication.place
London
ethz.publication.status
published
en_US
ethz.leitzahl
ETH Zürich::00002 - ETH Zürich::00012 - Lehre und Forschung::00007 - Departemente::02030 - Dep. Biologie / Dep. of Biology::02521 - Inst. f. Molekularbiologie u. Biophysik / Inst. Molecular Biology and Biophysics::09529 - Korkhov, Volodymyr / Korkhov, Volodymyr
ethz.leitzahl.certified
ETH Zürich::00002 - ETH Zürich::00012 - Lehre und Forschung::00007 - Departemente::02030 - Dep. Biologie / Dep. of Biology::02521 - Inst. f. Molekularbiologie u. Biophysik / Inst. Molecular Biology and Biophysics::09529 - Korkhov, Volodymyr / Korkhov, Volodymyr
ethz.grant.agreementno
166672
ethz.grant.fundername
SNF
ethz.grant.funderDoi
10.13039/501100001711
ethz.grant.program
Exzellenzbeitrag in Lebenswissenschaften
ethz.date.deposited
2017-06-12T19:44:21Z
ethz.source
ECIT
ethz.identifier.importid
imp593655405b57a66405
ethz.ecitpid
pub:191315
ethz.eth
yes
en_US
ethz.availability
Open access
en_US
ethz.rosetta.installDate
2017-07-18T15:42:36Z
ethz.rosetta.lastUpdated
2024-02-02T06:20:38Z
ethz.rosetta.versionExported
true
ethz.COinS
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