Live Cell Imaging of Enzymatic Turnover of an Adenosine 5′-Tetraphosphate Analog

Lade...
Vorschaubild
Dateien
Bhat_2-1p1ylvkh0d6j87.pdf
Bhat_2-1p1ylvkh0d6j87.pdfGröße: 2.14 MBDownloads: 107
Datum
2021
Herausgeber:innen
Kontakt
ISSN der Zeitschrift
Electronic ISSN
ISBN
Bibliografische Daten
Verlag
Schriftenreihe
Auflagebezeichnung
ArXiv-ID
Internationale Patentnummer
Link zur Lizenz
Angaben zur Forschungsförderung
Projekt
Open Access-Veröffentlichung
Open Access Gold
Core Facility der Universität Konstanz
Gesperrt bis
Titel in einer weiteren Sprache
Forschungsvorhaben
Organisationseinheiten
Zeitschriftenheft
Publikationstyp
Zeitschriftenartikel
Publikationsstatus
Published
Erschienen in
Zusammenfassung

The hydrolysis of nucleotides is of paramount importance as an energy source for cellular processes. In addition, the transfer of phosphates from nucleotides onto proteins is important as a post-translational protein modification. Monitoring the enzymatic turnover of nucleotides therefore offers great potential as a tool to follow enzymatic activity. While a number of fluorescence sensors are known, so far, there are no methods available for the real-time monitoring of ATP hydrolysis inside live cells. We present the synthesis and application of a novel fluorogenic adenosine 5′-tetraphosphate (Ap4) analog suited for this task. Upon enzymatic hydrolysis, the molecule displays an increase in fluorescence intensity, which provides a readout of its turnover. We demonstrate how this can be used for monitoring cellular processes involving Ap4 hydrolysis. To this end, we visualized the enzymatic activity in live cells using confocal fluorescence microscopy of the Ap4 analog. Our results demonstrate that the Ap4 analog is hydrolyzed in lysosomes. We show that this approach is suited to visualize the lysosome distribution profiles within the live cell and discuss how it can be employed to gather information regarding autophagic flux.

Zusammenfassung in einer weiteren Sprache
Fachgebiet (DDC)
540 Chemie
Schlagwörter
Konferenz
Rezension
undefined / . - undefined, undefined
Zitieren
ISO 690BHAT, Anayat, Shuang LI, Daniel HAMMLER, Martin WINTERHALDER, Andreas MARX, Andreas ZUMBUSCH, 2021. Live Cell Imaging of Enzymatic Turnover of an Adenosine 5′-Tetraphosphate Analog. In: International Journal of Molecular Sciences (IJMS). MDPI. 2021, 22(16), 8616. ISSN 1661-6596. eISSN 1422-0067. Available under: doi: 10.3390/ijms22168616
BibTex
@article{Bhat2021-08-10Imagi-54850,
  year={2021},
  doi={10.3390/ijms22168616},
  title={Live Cell Imaging of Enzymatic Turnover of an Adenosine 5′-Tetraphosphate Analog},
  number={16},
  volume={22},
  issn={1661-6596},
  journal={International Journal of Molecular Sciences (IJMS)},
  author={Bhat, Anayat and Li, Shuang and Hammler, Daniel and Winterhalder, Martin and Marx, Andreas and Zumbusch, Andreas},
  note={Article Number: 8616}
}
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/123456789/54850">
    <dc:contributor>Li, Shuang</dc:contributor>
    <dc:creator>Hammler, Daniel</dc:creator>
    <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2021-09-10T08:24:46Z</dc:date>
    <void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/>
    <dc:rights>Attribution 4.0 International</dc:rights>
    <dc:creator>Li, Shuang</dc:creator>
    <dc:contributor>Zumbusch, Andreas</dc:contributor>
    <dc:contributor>Marx, Andreas</dc:contributor>
    <dcterms:rights rdf:resource="http://creativecommons.org/licenses/by/4.0/"/>
    <dc:creator>Zumbusch, Andreas</dc:creator>
    <dc:creator>Marx, Andreas</dc:creator>
    <dcterms:issued>2021-08-10</dcterms:issued>
    <bibo:uri rdf:resource="https://kops.uni-konstanz.de/handle/123456789/54850"/>
    <dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/54850/1/Bhat_2-1p1ylvkh0d6j87.pdf"/>
    <dcterms:title>Live Cell Imaging of Enzymatic Turnover of an Adenosine 5′-Tetraphosphate Analog</dcterms:title>
    <dc:creator>Bhat, Anayat</dc:creator>
    <foaf:homepage rdf:resource="http://localhost:8080/"/>
    <dc:contributor>Winterhalder, Martin</dc:contributor>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
    <dc:contributor>Bhat, Anayat</dc:contributor>
    <dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2021-09-10T08:24:46Z</dcterms:available>
    <dcterms:abstract xml:lang="eng">The hydrolysis of nucleotides is of paramount importance as an energy source for cellular processes. In addition, the transfer of phosphates from nucleotides onto proteins is important as a post-translational protein modification. Monitoring the enzymatic turnover of nucleotides therefore offers great potential as a tool to follow enzymatic activity. While a number of fluorescence sensors are known, so far, there are no methods available for the real-time monitoring of ATP hydrolysis inside live cells. We present the synthesis and application of a novel fluorogenic adenosine 5′-tetraphosphate (Ap4) analog suited for this task. Upon enzymatic hydrolysis, the molecule displays an increase in fluorescence intensity, which provides a readout of its turnover. We demonstrate how this can be used for monitoring cellular processes involving Ap4 hydrolysis. To this end, we visualized the enzymatic activity in live cells using confocal fluorescence microscopy of the Ap4 analog. Our results demonstrate that the Ap4 analog is hydrolyzed in lysosomes. We show that this approach is suited to visualize the lysosome distribution profiles within the live cell and discuss how it can be employed to gather information regarding autophagic flux.</dcterms:abstract>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
    <dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/54850/1/Bhat_2-1p1ylvkh0d6j87.pdf"/>
    <dc:contributor>Hammler, Daniel</dc:contributor>
    <dc:language>eng</dc:language>
    <dc:creator>Winterhalder, Martin</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
Ja
Diese Publikation teilen