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  <doc>
    <id>62821</id>
    <completedYear/>
    <publishedYear>2025</publishedYear>
    <thesisYearAccepted/>
    <language>eng</language>
    <pageFirst>1</pageFirst>
    <pageLast>14</pageLast>
    <pageNumber/>
    <edition/>
    <issue/>
    <volume/>
    <type>preprint</type>
    <publisherName/>
    <publisherPlace/>
    <creatingCorporation/>
    <contributingCorporation/>
    <belongsToBibliography>0</belongsToBibliography>
    <completedDate>--</completedDate>
    <publishedDate>--</publishedDate>
    <thesisDateAccepted>--</thesisDateAccepted>
    <title language="eng">Yeast [FeFe]-hydrogenase-like protein Nar1 can bind not only two [4Fe-4S] clusters but also a [2Fe-2S] cluster</title>
    <abstract language="eng">Nar1 is an essential eukaryotic protein proposed to function as an iron-sulfur (Fe/S) cluster trafficking factor in the cytosolic iron-sulfur assembly (CIA) machinery. However, such a role has remained unclear due to difficulties in purifying adequate amounts of cofactor-bound protein. The [FeFe]-hydrogenase-like protein has two conserved binding sites for [4Fe-4S] clusters, one of which is predicted to be a labile site for cluster transfer to downstream targets. Here, we report a new preparation procedure for Nar1 that facilitated studies by UV-Vis, EPR, and Mössbauer spectroscopies, along with native mass spectrometry. Nar1 recombinantly produced in E. coli contained a [4Fe-4S] cluster, bound presumably at site 1, along with an unexpected [2Fe-2S] cluster bound at an unknown site. Fe/S reconstitution reactions installed a second [4Fe-4S] cluster at site 2, leading to protein with three Fe/S cofactors. Strikingly, one [4Fe-4S] cluster was rapidly destroyed by molecular oxygen, potentially linking Nar1 oxygen sensitivity to phenotypes observed previously in vivo. These advances now allow for the pursuit of in vitro Fe/S cluster transfer assays, which will shed light on Fe/S trafficking by CIA components and how they may facilitate the insertion of [4Fe-4S] and potentially [2Fe-2S] clusters into target proteins in the cytosol.</abstract>
    <parentTitle language="eng">bioRxiv</parentTitle>
    <identifier type="doi">10.1101/2025.03.25.644927</identifier>
    <identifier type="urn">urn:nbn:de:kobv:b43-628217</identifier>
    <enrichment key="opus.source">publish</enrichment>
    <enrichment key="opus.doi.autoCreate">false</enrichment>
    <enrichment key="opus.urn.autoCreate">true</enrichment>
    <licence>Creative Commons - CC BY - Namensnennung 4.0 International</licence>
    <author>Joseph J. Braymer</author>
    <author>L. Knauer</author>
    <author>J. C. Crack</author>
    <author>J. Oltmanns</author>
    <author>M. Heghmanns</author>
    <author>J. C. Soares</author>
    <author>N. E. Le Brun</author>
    <author>V. Schünemann</author>
    <author>M. Kasanmascheff</author>
    <subject>
      <language>eng</language>
      <type>uncontrolled</type>
      <value>Iron sulfur cluster</value>
    </subject>
    <subject>
      <language>eng</language>
      <type>uncontrolled</type>
      <value>Hydrogenase-like proteins</value>
    </subject>
    <subject>
      <language>eng</language>
      <type>uncontrolled</type>
      <value>Metalloprotein</value>
    </subject>
    <collection role="ddc" number="628">Sanitär- und Kommunaltechnik; Umwelttechnik</collection>
    <collection role="institutes" number="">1 Analytische Chemie; Referenzmaterialien</collection>
    <collection role="institutes" number="">1.8 Umweltanalytik</collection>
    <collection role="themenfelder" number="">Umwelt</collection>
    <collection role="fulltextaccess" number="">Datei für die Öffentlichkeit verfügbar ("Open Access")</collection>
    <collection role="literaturgattung" number="">Graue Literatur</collection>
    <collection role="themenfelder" number="">Biokorrosion</collection>
    <collection role="unnumberedseries" number="">Preprints der BAM</collection>
    <thesisPublisher>Bundesanstalt für Materialforschung und -prüfung (BAM)</thesisPublisher>
    <file>https://opus4.kobv.de/opus4-bam/files/62821/Nar1-FeS-Char_MS_bioRxiv.pdf</file>
  </doc>
</export-example>
