PH-dependent conformational changes of KcsA tetramer and monomer probed by Raman spectroscopy

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  • Helmholtz Zentrum München - Deutsches Forschungszentrum für Gesundheit und Umwelt
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OriginalspracheEnglisch
Aufsatznummer2736
FachzeitschriftInternational Journal of Molecular Sciences
Jahrgang20
Ausgabenummer11
PublikationsstatusVeröffentlicht - 1 Juni 2019

Abstract

KcsA is a tetrameric potassium channel formed out of four identical monomeric subunits used as a standard model for selective potassium transport and pH-dependent gating. Large conformational changes are reported for tetramer and monomer upon gating, and the response of the monomer being controversial with the two major studies partially contradicting each other. KcsA was analyzed as functional tetramers embedded in liposomes and as monomer subunits with confocal Raman microscopy under physiological conditions for the active and the closed channel state, using 532 nm excitation to avoid introducing conformational changes during the measurement. Channel function was confirmed using liposome flux assay. While the classic fingerprint region below 1800 rel. cm−1 in the Raman spectrum of the tetramer was unaffected, the CH-stretching region between 2800 and 3200 rel. cm−1 was found to be strongly affected by the conformation. No pH-dependency was observed in the Raman spectra of the monomer subunits, which closely resembled the Raman spectrum of the tetramer in its active conformation, indicating that the open conformation of the monomer and not the closed conformation as postulated may equal the relaxed state of the molecule.

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PH-dependent conformational changes of KcsA tetramer and monomer probed by Raman spectroscopy. / Kniggendorf, Ann Kathrin; Schmidt, David; Roth, Bernhard et al.
in: International Journal of Molecular Sciences, Jahrgang 20, Nr. 11, 2736, 01.06.2019.

Publikation: Beitrag in FachzeitschriftArtikelForschungPeer-Review

Kniggendorf AK, Schmidt D, Roth B, Plettenburg O, Zeilinger C. PH-dependent conformational changes of KcsA tetramer and monomer probed by Raman spectroscopy. International Journal of Molecular Sciences. 2019 Jun 1;20(11):2736. doi: 10.3390/ijms20112736, 10.15488/5102
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title = "PH-dependent conformational changes of KcsA tetramer and monomer probed by Raman spectroscopy",
abstract = "KcsA is a tetrameric potassium channel formed out of four identical monomeric subunits used as a standard model for selective potassium transport and pH-dependent gating. Large conformational changes are reported for tetramer and monomer upon gating, and the response of the monomer being controversial with the two major studies partially contradicting each other. KcsA was analyzed as functional tetramers embedded in liposomes and as monomer subunits with confocal Raman microscopy under physiological conditions for the active and the closed channel state, using 532 nm excitation to avoid introducing conformational changes during the measurement. Channel function was confirmed using liposome flux assay. While the classic fingerprint region below 1800 rel. cm−1 in the Raman spectrum of the tetramer was unaffected, the CH-stretching region between 2800 and 3200 rel. cm−1 was found to be strongly affected by the conformation. No pH-dependency was observed in the Raman spectra of the monomer subunits, which closely resembled the Raman spectrum of the tetramer in its active conformation, indicating that the open conformation of the monomer and not the closed conformation as postulated may equal the relaxed state of the molecule.",
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AU - Kniggendorf, Ann Kathrin

AU - Schmidt, David

AU - Roth, Bernhard

AU - Plettenburg, Oliver

AU - Zeilinger, Carsten

N1 - Funding information: A.-K.K. received funding by the German Bundesministerium für Bildung und Forschung (BMBF, Federal Ministry of Education and Research) within the collaborative project OPTIMUS (13N13811) and by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation)—Project ID 397827619. B.R. was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Germany’s Excellence Strategy within the Cluster of Excellence PhoenixD (EXC 2122, Project ID 390833453). The publication of this article was funded by the Open Access Fund of the Leibniz Universität Hannover.

PY - 2019/6/1

Y1 - 2019/6/1

N2 - KcsA is a tetrameric potassium channel formed out of four identical monomeric subunits used as a standard model for selective potassium transport and pH-dependent gating. Large conformational changes are reported for tetramer and monomer upon gating, and the response of the monomer being controversial with the two major studies partially contradicting each other. KcsA was analyzed as functional tetramers embedded in liposomes and as monomer subunits with confocal Raman microscopy under physiological conditions for the active and the closed channel state, using 532 nm excitation to avoid introducing conformational changes during the measurement. Channel function was confirmed using liposome flux assay. While the classic fingerprint region below 1800 rel. cm−1 in the Raman spectrum of the tetramer was unaffected, the CH-stretching region between 2800 and 3200 rel. cm−1 was found to be strongly affected by the conformation. No pH-dependency was observed in the Raman spectra of the monomer subunits, which closely resembled the Raman spectrum of the tetramer in its active conformation, indicating that the open conformation of the monomer and not the closed conformation as postulated may equal the relaxed state of the molecule.

AB - KcsA is a tetrameric potassium channel formed out of four identical monomeric subunits used as a standard model for selective potassium transport and pH-dependent gating. Large conformational changes are reported for tetramer and monomer upon gating, and the response of the monomer being controversial with the two major studies partially contradicting each other. KcsA was analyzed as functional tetramers embedded in liposomes and as monomer subunits with confocal Raman microscopy under physiological conditions for the active and the closed channel state, using 532 nm excitation to avoid introducing conformational changes during the measurement. Channel function was confirmed using liposome flux assay. While the classic fingerprint region below 1800 rel. cm−1 in the Raman spectrum of the tetramer was unaffected, the CH-stretching region between 2800 and 3200 rel. cm−1 was found to be strongly affected by the conformation. No pH-dependency was observed in the Raman spectra of the monomer subunits, which closely resembled the Raman spectrum of the tetramer in its active conformation, indicating that the open conformation of the monomer and not the closed conformation as postulated may equal the relaxed state of the molecule.

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KW - KcsA

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