Gd(III)-PyMTA Label Is Suitable for In-Cell EPR

Lade...
Vorschaubild
Dateien
Qi_2-p9hn4pcv4zkf3.pdf
Qi_2-p9hn4pcv4zkf3.pdfGröße: 464.29 KBDownloads: 25
Datum
2014
Autor:innen
Herausgeber:innen
Kontakt
ISSN der Zeitschrift
Electronic ISSN
ISBN
Bibliografische Daten
Verlag
Schriftenreihe
Auflagebezeichnung
DOI (zitierfähiger Link)
ArXiv-ID
Internationale Patentnummer
Link zur Lizenz
oops
Angaben zur Forschungsförderung
Projekt
Open Access-Veröffentlichung
Open Access Green
Core Facility der Universität Konstanz
Gesperrt bis
Titel in einer weiteren Sprache
Forschungsvorhaben
Organisationseinheiten
Zeitschriftenheft
Publikationstyp
Zeitschriftenartikel
Publikationsstatus
Published
Erschienen in
Zusammenfassung

Distance measurement in the nanometer range by electron paramagnetic resonance spectroscopy (EPR) in combination with site-directed spin labeling is a very powerful tool to monitor the structure and dynamics of biomacromolecules in their natural environment. However, in-cell application is hampered by the short lifetime of the commonly used nitroxide spin labels in the reducing milieu inside a cell. Here, we demonstrate that the Gd(III) based spin label Gd-PyMTA is suitable for in-cell EPR. Gd-PyMTA turned out to be cell compatible and was proven to be inert in in-cell extracts of Xenopus laevis oocytes at 18 °C for more than 24 h. The proline rich peptide H-AP10CP10CP10-NH2 was site-directedly spin labeled with Gd-PyMTA at both cysteine moieties. The resulting peptide, H-AP10C(Gd-PyMTA)P10C(Gd-PyMTA)P10-NH2, as well as the model compound Gd-spacer-Gd, which consists of a spacer of well-known stiffness, were microinjected into Xenopus laevis oocytes, and the Gd(III)–Gd(III) distances were determined by double electron–electron resonance (DEER) spectroscopy. To analyze the intracellular peptide conformation, a rotamer library was set up to take the conformational flexibility of the tether between the Gd(III) ion and the Cα of the cysteine moiety into account. The results suggest that the spin labeled peptide H-AP10C(Gd-PyMTA)P10C(Gd-PyMTA)P10-NH2 is inserted into cell membranes, coinciding with a conformational change of the oligoproline from a PPII into a PPI helix.

Zusammenfassung in einer weiteren Sprache
Fachgebiet (DDC)
540 Chemie
Schlagwörter
Konferenz
Rezension
undefined / . - undefined, undefined
Zitieren
ISO 690QI, Mian, Andreas GROSS, Gunnar JESCHKE, Adelheid GODT, Malte DRESCHER, 2014. Gd(III)-PyMTA Label Is Suitable for In-Cell EPR. In: Journal of the American Chemical Society. 2014, 136(43), pp. 15366-15378. ISSN 0002-7863. eISSN 1520-5126. Available under: doi: 10.1021/ja508274d
BibTex
@article{Qi2014GdIII-30104,
  year={2014},
  doi={10.1021/ja508274d},
  title={Gd(III)-PyMTA Label Is Suitable for In-Cell EPR},
  number={43},
  volume={136},
  issn={0002-7863},
  journal={Journal of the American Chemical Society},
  pages={15366--15378},
  author={Qi, Mian and Groß, Andreas and Jeschke, Gunnar and Godt, Adelheid and Drescher, Malte}
}
RDF
<rdf:RDF
    xmlns:dcterms="http://purl.org/dc/terms/"
    xmlns:dc="http://purl.org/dc/elements/1.1/"
    xmlns:rdf="http://www.w3.org/1999/02/22-rdf-syntax-ns#"
    xmlns:bibo="http://purl.org/ontology/bibo/"
    xmlns:dspace="http://digital-repositories.org/ontologies/dspace/0.1.0#"
    xmlns:foaf="http://xmlns.com/foaf/0.1/"
    xmlns:void="http://rdfs.org/ns/void#"
    xmlns:xsd="http://www.w3.org/2001/XMLSchema#" > 
  <rdf:Description rdf:about="https://kops.uni-konstanz.de/server/rdf/resource/123456789/30104">
    <dc:creator>Groß, Andreas</dc:creator>
    <dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2015-02-27T10:34:03Z</dcterms:available>
    <dcterms:issued>2014</dcterms:issued>
    <void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/>
    <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2015-02-27T10:34:03Z</dc:date>
    <dc:contributor>Drescher, Malte</dc:contributor>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
    <dc:language>eng</dc:language>
    <dc:contributor>Groß, Andreas</dc:contributor>
    <dc:contributor>Qi, Mian</dc:contributor>
    <dc:contributor>Godt, Adelheid</dc:contributor>
    <bibo:uri rdf:resource="http://kops.uni-konstanz.de/handle/123456789/30104"/>
    <foaf:homepage rdf:resource="http://localhost:8080/"/>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/52"/>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
    <dc:creator>Jeschke, Gunnar</dc:creator>
    <dcterms:abstract xml:lang="eng">Distance measurement in the nanometer range by electron paramagnetic resonance spectroscopy (EPR) in combination with site-directed spin labeling is a very powerful tool to monitor the structure and dynamics of biomacromolecules in their natural environment. However, in-cell application is hampered by the short lifetime of the commonly used nitroxide spin labels in the reducing milieu inside a cell. Here, we demonstrate that the Gd(III) based spin label Gd-PyMTA is suitable for in-cell EPR. Gd-PyMTA turned out to be cell compatible and was proven to be inert in in-cell extracts of Xenopus laevis oocytes at 18 °C for more than 24 h. The proline rich peptide H-AP10CP&lt;sub&gt;10&lt;/sub&gt;CP&lt;sub&gt;10&lt;/sub&gt;-NH&lt;sub&gt;2&lt;/sub&gt; was site-directedly spin labeled with Gd-PyMTA at both cysteine moieties. The resulting peptide, H-AP&lt;sub&gt;10&lt;/sub&gt;C(Gd-PyMTA)P&lt;sub&gt;10&lt;/sub&gt;C(Gd-PyMTA)P&lt;sub&gt;10&lt;/sub&gt;-NH&lt;sub&gt;2&lt;/sub&gt;, as well as the model compound Gd-spacer-Gd, which consists of a spacer of well-known stiffness, were microinjected into Xenopus laevis oocytes, and the Gd(III)–Gd(III) distances were determined by double electron–electron resonance (DEER) spectroscopy. To analyze the intracellular peptide conformation, a rotamer library was set up to take the conformational flexibility of the tether between the Gd(III) ion and the C&lt;sub&gt;α&lt;/sub&gt; of the cysteine moiety into account. The results suggest that the spin labeled peptide H-AP&lt;sub&gt;10&lt;/sub&gt;C(Gd-PyMTA)P&lt;sub&gt;10&lt;/sub&gt;C(Gd-PyMTA)P&lt;sub&gt;10&lt;/sub&gt;-NH&lt;sub&gt;2&lt;/sub&gt; is inserted into cell membranes, coinciding with a conformational change of the oligoproline from a PPII into a PPI helix.</dcterms:abstract>
    <dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/30104/1/Qi_2-p9hn4pcv4zkf3.pdf"/>
    <dc:contributor>Jeschke, Gunnar</dc:contributor>
    <dcterms:title>Gd(III)-PyMTA Label Is Suitable for In-Cell EPR</dcterms:title>
    <dc:creator>Godt, Adelheid</dc:creator>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/52"/>
    <dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/30104/1/Qi_2-p9hn4pcv4zkf3.pdf"/>
    <dc:creator>Qi, Mian</dc:creator>
    <dc:creator>Drescher, Malte</dc:creator>
  </rdf:Description>
</rdf:RDF>
Interner Vermerk
xmlui.Submission.submit.DescribeStep.inputForms.label.kops_note_fromSubmitter
Kontakt
URL der Originalveröffentl.
Prüfdatum der URL
Prüfungsdatum der Dissertation
Finanzierungsart
Kommentar zur Publikation
Allianzlizenz
Corresponding Authors der Uni Konstanz vorhanden
Internationale Co-Autor:innen
Universitätsbibliographie
Ja
Begutachtet
Diese Publikation teilen