Publikation: Structural and catalytic insights into HoLaMa, a derivative of Klenow DNA polymerase lacking the proofreading domain
Lade...
Dateien
Datum
2019
Autor:innen
Herausgeber:innen
ISSN der Zeitschrift
Electronic ISSN
ISBN
Bibliografische Daten
Verlag
Schriftenreihe
Auflagebezeichnung
URI (zitierfähiger Link)
DOI (zitierfähiger Link)
Internationale Patentnummer
Link zur Lizenz
Angaben zur Forschungsförderung
Projekt
Open Access-Veröffentlichung
Open Access Gold
Sammlungen
Core Facility der Universität Konstanz
Titel in einer weiteren Sprache
Publikationstyp
Zeitschriftenartikel
Publikationsstatus
Published
Erschienen in
PLoS one. 2019, 14(4), e0215411. eISSN 1932-6203. Available under: doi: 10.1371/journal.pone.0215411
Zusammenfassung
We report here on the stability and catalytic properties of the HoLaMa DNA polymerase, a Klenow sub-fragment lacking the 3’-5’ exonuclease domain. HoLaMa was overexpressed in Escherichia coli, and the enzyme was purified by means of standard chromatographic techniques. High-resolution NMR experiments revealed that HoLaMa is properly folded at pH 8.0 and 20°C. In addition, urea induced a cooperative folding to unfolding transition of HoLaMa, possessing an overall thermodynamic stability and a transition midpoint featuring ΔG and CM equal to (15.7 ± 1.9) kJ/mol and (3.5 ± 0.6) M, respectively. When the catalytic performances of HoLaMa were compared to those featured by the Klenow enzyme, we did observe a 10-fold lower catalytic efficiency by the HoLaMa enzyme. Surprisingly, HoLaMa and Klenow DNA polymerases possess markedly different sensitivities in competitive inhibition assays performed to test the effect of single dNTPs.
Zusammenfassung in einer weiteren Sprache
Fachgebiet (DDC)
540 Chemie
Schlagwörter
Konferenz
Rezension
undefined / . - undefined, undefined
Zitieren
ISO 690
KOVERMANN, Michael, Alessandra STEFAN, Anna CASTALDO, Sara CARAMIA, Alejandro HOCHKOEPPLER, 2019. Structural and catalytic insights into HoLaMa, a derivative of Klenow DNA polymerase lacking the proofreading domain. In: PLoS one. 2019, 14(4), e0215411. eISSN 1932-6203. Available under: doi: 10.1371/journal.pone.0215411BibTex
@article{Kovermann2019Struc-46067,
year={2019},
doi={10.1371/journal.pone.0215411},
title={Structural and catalytic insights into HoLaMa, a derivative of Klenow DNA polymerase lacking the proofreading domain},
number={4},
volume={14},
journal={PLoS one},
author={Kovermann, Michael and Stefan, Alessandra and Castaldo, Anna and Caramia, Sara and Hochkoeppler, Alejandro},
note={Article Number: e0215411}
}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/123456789/46067">
<dc:creator>Stefan, Alessandra</dc:creator>
<dcterms:abstract xml:lang="eng">We report here on the stability and catalytic properties of the HoLaMa DNA polymerase, a Klenow sub-fragment lacking the 3’-5’ exonuclease domain. HoLaMa was overexpressed in Escherichia coli, and the enzyme was purified by means of standard chromatographic techniques. High-resolution NMR experiments revealed that HoLaMa is properly folded at pH 8.0 and 20°C. In addition, urea induced a cooperative folding to unfolding transition of HoLaMa, possessing an overall thermodynamic stability and a transition midpoint featuring ΔG and C<sub>M</sub> equal to (15.7 ± 1.9) kJ/mol and (3.5 ± 0.6) M, respectively. When the catalytic performances of HoLaMa were compared to those featured by the Klenow enzyme, we did observe a 10-fold lower catalytic efficiency by the HoLaMa enzyme. Surprisingly, HoLaMa and Klenow DNA polymerases possess markedly different sensitivities in competitive inhibition assays performed to test the effect of single dNTPs.</dcterms:abstract>
<dcterms:title>Structural and catalytic insights into HoLaMa, a derivative of Klenow DNA polymerase lacking the proofreading domain</dcterms:title>
<dc:creator>Kovermann, Michael</dc:creator>
<dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2019-06-21T12:28:24Z</dcterms:available>
<foaf:homepage rdf:resource="http://localhost:8080/"/>
<dc:language>eng</dc:language>
<void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/>
<dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2019-06-21T12:28:24Z</dc:date>
<dc:contributor>Castaldo, Anna</dc:contributor>
<dcterms:rights rdf:resource="http://creativecommons.org/licenses/by/4.0/"/>
<dc:creator>Castaldo, Anna</dc:creator>
<dc:rights>Attribution 4.0 International</dc:rights>
<dc:creator>Hochkoeppler, Alejandro</dc:creator>
<dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
<dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/46067/1/Kovermann_2-1akcs347hhnhb4.pdf"/>
<dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/29"/>
<dc:creator>Caramia, Sara</dc:creator>
<dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/46067/1/Kovermann_2-1akcs347hhnhb4.pdf"/>
<dc:contributor>Stefan, Alessandra</dc:contributor>
<dcterms:issued>2019</dcterms:issued>
<dc:contributor>Caramia, Sara</dc:contributor>
<dc:contributor>Kovermann, Michael</dc:contributor>
<bibo:uri rdf:resource="https://kops.uni-konstanz.de/handle/123456789/46067"/>
<dc:contributor>Hochkoeppler, Alejandro</dc:contributor>
</rdf:Description>
</rdf:RDF>Interner Vermerk
xmlui.Submission.submit.DescribeStep.inputForms.label.kops_note_fromSubmitter
Prüfungsdatum der Dissertation
Finanzierungsart
Kommentar zur Publikation
Allianzlizenz
Corresponding Authors der Uni Konstanz vorhanden
Internationale Co-Autor:innen
Universitätsbibliographie
Ja
Begutachtet
Ja
