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ETH-Rektor Günther Dissertori überreicht Charlotte Bopp den Otto-Jaag-Gewässerschutzpreis 2023 (Foto: ETH, Giulia Marthaler)

Otto-Jaag-Gewässerschutzpreis 2023 und ETH-Medaille für Charlotte Bopp

21 novembre 2023 | Claudia Carle

Für ihre Dissertation wird die Umweltwissenschaftlerin Charlotte Bopp gleich doppelt ausgezeichnet: Am ETH-Tag vom 18. November erhielt sie den Otto-Jaag-Gewässerschutzpreis. Im Januar wird sie auch noch die ETH-Medaille entgegennehmen. Ihre Arbeit leistet einen wichtigen Beitrag zum Verständnis des biologischen Abbaus von organischen Schadstoffen in der Umwelt.

Mit dem Otto-Jaag-Gewässerschutzpreis zeichnet die ETH Zürich hervorragende Master- und Doktorarbeiten auf dem Gebiet des Gewässerschutzes und der Gewässerkunde aus. Charlotte Bopp durfte diese Auszeichnung am ETH-Tag vom 18. November für ihre Dissertation zum Thema «The role of oxygen uncoupling by Rieske non-heme iron dioxygenases in the biodegradation of aromatic contaminants” entgegennehmen. Im Januar wird ihr auch noch die ETH-Medaille verliehen, welche die ETH Zürich für herausragende Master- und Doktorarbeiten vergibt.

Ineffiziente Oxidation der Schadstoffe

Als Doktorandin in der Abteilung Umweltchemie des Wasserforschungsinstitutes Eawag nahm Charlotte Bopp den biologischen Abbau schwer abbaubarer, organischer Schadstoffe unter die Lupe. Gelangen solche aromatischen Verbindungen, die zum Beispiel in Pestiziden, Medikamenten oder Sprengstoffen vorkommen, in Böden und Gewässer, können Mikroorganismen diese dank einer Gruppe von Enzymen mit dem Namen «Rieske Oxygenasen» oxidieren und so abbauen. Bopp konzentrierte sich in ihrer Arbeit auf die Untergruppe der Sprengstoff-abbauenden Enzyme und wollte wissen, wie effizient diese arbeiten. Ihre Ergebnisse stellen den Enzymen kein gutes Zeugnis aus. Statt den Sauerstoff direkt auf die Schadstoffe zu übertragen, bilden die Enzyme zuerst eine besonders reaktive Form des Sauerstoffs. Nur rund die Hälfte davon reagiert anschliessend aber tatsächlich mit den Schadstoffen, der andere Teil des reaktionsfreudigen Sauerstoffs oxidiert alle möglichen anderen Substanzen in den Mikroorganismen. Das kann zum Nachteil der Mikroorganismen sein und diese schädigen.

Die Neuen sind effizienter

Aber dieser Prozess kann auch Vorteile mit sich bringen, wie Bopp zeigen konnte: Kommen die Mikroorganismen mit neuen Schadstoffen in Kontakt, für deren Abbau ihr bestehendes Enzymspektrum nicht geeignet ist, können sie sich anpassen. Der reaktive Sauerstoff führt bei den Enzymen zu punktuellen Mutationen, wodurch sich einzelne Aminosäuren innerhalb des Enzyms verändern und dadurch neue Enzyme entstehen. Einige davon arbeiten sogar effizienter als die ursprünglichen. Dank dieses evolutiven Prozesses können die Mikroorganismen nach einiger Zeit die neuen Schadstoffe verwerten.

«Mit ihrer Forschungsarbeit hat Charlotte Bopp Zusammenhänge beim biologischen Abbau von Schadstoffen aufgedeckt, die bisher nicht bekannt waren», sagt Thomas Hofstetter, Leiter der Abteilung Umweltchemie an der Eawag, der ihre Dissertation betreut hat. Bisher habe man rein auf Grund der Menge vorhandener Enzyme in der Umwelt auf die Kapazität zum Abbau von Schadstoffen geschlossen. «Die Resultate von Charlotte Bopp zeigen, dass man hier genauer hinschauen und die unterschiedliche Effizienz der Organismen und ihrer Enzyme mitberücksichtigen muss.»

Charlotte Bopp freut sich über die grosse Anerkennung ihrer Arbeit: «Wir haben uns getraut dahin zu schauen, wo diese Enzyme scheinbar versagen.» Dabei sei es genau die Fehlerhaftigkeit, die den Mikroorganismen auf lange Sicht erlaube mit vielfältigen Schadstoffen umzugehen. «Dieses Rätsel konnten wir nur aufklären dank der interdisziplinären Aufstellung der Eawag und einem Team, das als Ganzes diese Auszeichnungen verdient hat», erklärt Bopp. Seit Abschluss ihrer Dissertation arbeitet sie in der Industrie und beschäftigt sich mit der Weiterentwicklung von Ozonierungsprozessen in der Trink- und Abwasserbehandlung.
 

Titelbild: ETH-Rektor Günther Dissertori überreicht Charlotte Bopp den Otto-Jaag-Gewässerschutzpreis 2023 (Foto: ETH, Giulia Marthaler)
 

Originalpublikationen

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      title => protected'Elucidating the role of O<sub>2</sub> uncoupling in the oxidative biodegrada
         tion of organic contaminants by Rieske non-heme iron dioxygenases' (141 chars)
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      categories => protected'non-heme ferrous iron oxygenases; Rieske oxygenases; biocatalysis; O2 uncoup
         ling; O2 activation; kinetic isotope effect; biodegradation' (135 chars)
      description => protected'Oxygenations of aromatic soil and water contaminants with molecular O<sub>2<
         /sub> catalyzed by Rieske dioxygenases are frequent initial steps of biodegr
         adation in natural and engineered environments. Many of these non-heme ferro
         us iron enzymes are known to be involved in contaminant metabolism, but the 
         understanding of enzyme-substrate interactions that lead to successful biode
         gradation is still elusive. Here, we studied the mechanisms of O<sub>2</sub>
          activation and substrate hydroxylation of two nitroarene dioxygenases to ev
         aluate enzyme- and substrate-specific factors that determine the efficiency 
         of oxygenated product formation. Experiments in enzyme assays of 2-nitrotolu
         ene dioxygenase (2NTDO) and nitrobenzene dioxygenase (NBDO) with methyl-, fl
         uoro-, chloro-, and hydroxy-substituted nitroaromatic substrates reveal that
          typically 20-100% of the enzyme's activity involves unproductive paths of O
         <sub>2</sub> activation with generation of reactive oxygen species through s
         o-called O<sub>2</sub> uncoupling. The <sup>18</sup>O and <sup>13</sup>C kin
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          hydroxylation, respectively, suggest that O<sub>2</sub> uncoupling occurs a
         fter generation of Fe<sup>III</sup>-(hydro)peroxo species in the catalytic c
         ycle. While 2NTDO hydroxylates <em>ortho</em>-substituted nitroaromatic subs
         trates more efficiently, NBDO favors <em>meta</em>-substituted, presumably d
         ue to distinct active site residues of the two enzymes. Our data implies, ho
         wever, that the O<sub>2</sub> uncoupling and hydroxylation activity cannot b
         e assessed from simple structure-reactivity relationships. By quantifying O<
         sub>2</sub> uncoupling by Rieske dioxygenases, our work provides a mechanist
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      description => protected'Rieske dioxygenases catalyze the initial steps in the hydroxylation of aroma
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         se capable of transforming nitroarenes to nitrite and substituted catechols,
          that unproductive O<sub>2</sub> activation with the release of the unreacte
         d substrate and reactive oxygen species represents an important path in the 
         catalytic cycle. Through correlation of O<sub>2</sub> uncoupling for a serie
         s of substituted nitroaromatic compounds with <sup>18</sup>O and <sup>13</su
         p>C kinetic isotope effects of dissolved O<sub>2</sub> and aromatic substrat
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         e-limiting formation of Fe<sup>III</sup>-(hydro)peroxo species from which su
         bstrates are hydroxylated. Substituent effects on the extent of O<sub>2</sub
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Elucidating the role of O2 uncoupling in the oxidative biodegradation of organic contaminants by Rieske non-heme iron dioxygenases

Oxygenations of aromatic soil and water contaminants with molecular O2 catalyzed by Rieske dioxygenases are frequent initial steps of biodegradation in natural and engineered environments. Many of these non-heme ferrous iron enzymes are known to be involved in contaminant metabolism, but the understanding of enzyme-substrate interactions that lead to successful biodegradation is still elusive. Here, we studied the mechanisms of O2 activation and substrate hydroxylation of two nitroarene dioxygenases to evaluate enzyme- and substrate-specific factors that determine the efficiency of oxygenated product formation. Experiments in enzyme assays of 2-nitrotoluene dioxygenase (2NTDO) and nitrobenzene dioxygenase (NBDO) with methyl-, fluoro-, chloro-, and hydroxy-substituted nitroaromatic substrates reveal that typically 20-100% of the enzyme's activity involves unproductive paths of O2 activation with generation of reactive oxygen species through so-called O2 uncoupling. The 18O and 13C kinetic isotope effects of O2 activation and nitroaromatic substrate hydroxylation, respectively, suggest that O2 uncoupling occurs after generation of FeIII-(hydro)peroxo species in the catalytic cycle. While 2NTDO hydroxylates ortho-substituted nitroaromatic substrates more efficiently, NBDO favors meta-substituted, presumably due to distinct active site residues of the two enzymes. Our data implies, however, that the O2 uncoupling and hydroxylation activity cannot be assessed from simple structure-reactivity relationships. By quantifying O2 uncoupling by Rieske dioxygenases, our work provides a mechanistic link between contaminant biodegradation, the generation of reactive oxygen species, and possible adaptation strategies of microorganisms to the exposure of new contaminants.
Bopp, C. E.; Bernet, N. M.; Kohler, H.-P. E.; Hofstetter, T. B. (2022) Elucidating the role of O2 uncoupling in the oxidative biodegradation of organic contaminants by Rieske non-heme iron dioxygenases, ACS Environmental Au, 2(5), 428-440, doi:10.1021/acsenvironau.2c00023, Institutional Repository

Substrate-specific coupling of O2 activation to hydroxylations of aromatic compounds by rieske non-heme iron dioxygenases

Rieske dioxygenases catalyze the initial steps in the hydroxylation of aromatic compounds and are critical for the metabolism of xenobiotic substances. Because substrates do not bind to the mononuclear non-heme FeII center, elementary steps leading to O2 activation and substrate hydroxylation are difficult to delineate, thus making it challenging to rationalize divergent observations on enzyme mechanisms, reactivity, and substrate specificity. Here, we show for nitrobenzene dioxygenase, a Rieske dioxygenase capable of transforming nitroarenes to nitrite and substituted catechols, that unproductive O2 activation with the release of the unreacted substrate and reactive oxygen species represents an important path in the catalytic cycle. Through correlation of O2 uncoupling for a series of substituted nitroaromatic compounds with 18O and 13C kinetic isotope effects of dissolved O2 and aromatic substrates, respectively, we show that O2 uncoupling occurs after the rate-limiting formation of FeIII-(hydro)peroxo species from which substrates are hydroxylated. Substituent effects on the extent of O2 uncoupling suggest that the positioning of the substrate in the active site rather than the susceptibility of the substrate for attack by electrophilic oxygen species is responsible for unproductive O2 uncoupling. The proposed catalytic cycle provides a mechanistic basis for assessing the very different efficiencies of substrate hydroxylation vs unproductive O2 activation and generation of reactive oxygen species in reactions catalyzed by Rieske dioxygenases.
Pati, S. G.; Bopp, C. E.; Kohler, H.-P. E.; Hofstetter, T. B. (2022) Substrate-specific coupling of O2 activation to hydroxylations of aromatic compounds by rieske non-heme iron dioxygenases, ACS Catalysis, 12(11), 6444-6456, doi:10.1021/acscatal.2c00383, Institutional Repository

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