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dc.contributor.authorGrgic, Miriam
dc.contributor.authorWilliamson, Adele Kim
dc.contributor.authorBjerga, Gro Elin Kjæreng
dc.contributor.authorAltermark, Bjørn
dc.contributor.authorLeiros, Ingar
dc.date.accessioned2019-01-11T09:39:53Z
dc.date.available2019-01-11T09:39:53Z
dc.date.issued2018-05-25
dc.description.abstract<p>Cytosine-specific DNA methyltransferases are important enzymes in most living organisms. In prokaryotes, most DNA methyltransferases are members of the type II restriction-modification system where they methylate host DNA, thereby protecting it from digestion by the accompanying restriction endonucleases. DNA methyltransferases can also act as solitary enzymes having important roles in controlling gene expression, DNA replication, cell cycle and DNA post-replicative mismatch repair. They have potential applications in biotechnology, such as in labeling of biopolymers, DNA mapping or epigenetic analysis, as well as for general DNA-protein interaction studies.</p> <p>The <i>parI</i> gene from the psychrophilic bacterium <i>Psychrobacter</i> arcticus 273–4 encodes a cytosine-specific DNA methyltransferase. In this work, recombinant ParI was expressed and purified in fusion to either an N-terminal hexahistidine affinity tag, or a maltose binding protein following the hexahistidine affinity tag, for solubility improvement. After removal of the fusion partners, recombinant ParI was found to be monomeric by size exclusion chromatography, with its molecular mass estimated to be 54 kDa. The apparent melting temperature of the protein was 53 °C with no detectable secondary structures above 65 °C. Both recombinant and native ParI showed methyltransferase activity <i>in vivo</i>. In addition, MBP- and His-tagged ParI also demonstrated <i>in vitro</i> activity. Although the overall structure of ParI exhibits high thermal stability, the loss of <i>in vitro</i> activity upon removal of solubility tags or purification from the cellular milieu indicates that the catalytically active form is more labile. Horizontal gene transfer may explain the acquisition of a protein-encoding gene that does not display common cold-adapted features.</p>en_US
dc.description.sponsorshipUiT The Arctic University of Norwayen_US
dc.descriptionAccepted manuscript version. Published version available at <a href=https://doi.org/10.1016/j.pep.2018.05.012> https://doi.org/10.1016/j.pep.2018.05.012</a>.en_US
dc.identifier.citationGrgic, M., Williamson, A.K., Bjerga, G.E., Altermark, B. & Leiros, I. (2018). Biochemical characterization of ParI, an orphan C5-DNA methyltransferase from Psychrobacter arcticus 273-4. <i>Protein Expression and Purification</i>, 150, 100-108. https://doi.org/10.1016/j.pep.2018.05.012en_US
dc.identifier.cristinIDFRIDAID 1588003
dc.identifier.doi10.1016/j.pep.2018.05.012
dc.identifier.issn1046-5928
dc.identifier.issn1096-0279
dc.identifier.urihttps://hdl.handle.net/10037/14420
dc.language.isoengen_US
dc.publisherElsevieren_US
dc.relation.journalProtein Expression and Purification
dc.relation.projectIDinfo:eu-repo/grantAgreement/RCN/BIOTEK2021/244247/Norway/Engineering efficient DNA ligases for improved Next-Generation-Sequencing//en_US
dc.rights.accessRightsopenAccessen_US
dc.subjectVDP::Mathematics and natural science: 400::Chemistry: 440en_US
dc.subjectVDP::Matematikk og Naturvitenskap: 400::Kjemi: 440en_US
dc.subjectDNA methylationen_US
dc.subjectDNA methyltransferaseen_US
dc.subjectParIen_US
dc.subjectPsychrobacter arcticus 273–4en_US
dc.subjectPhage originen_US
dc.titleBiochemical characterization of ParI, an orphan C5-DNA methyltransferase from Psychrobacter arcticus 273-4en_US
dc.typeJournal articleen_US
dc.typeTidsskriftartikkelen_US
dc.typePeer revieweden_US


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