Preprints
https://doi.org/10.5194/egusphere-2025-1193
https://doi.org/10.5194/egusphere-2025-1193
18 Mar 2025
 | 18 Mar 2025

Variability in oxygen isotopic fractionation of enzymatic O2 consumption

Carolina F.M. de Carvalho, Moritz F. Lehmann, and Sarah G. Pati

Abstract. Stable isotope analysis of O2 has emerged as a valuable tool to study O2 dynamics at various environmental scales, from molecular mechanisms to ecosystem processes. Despite its utility, there is a lack of fundamental understanding of the large variability observed in O2 isotopic fractionation at the environment- and even enzyme-level. To expand our knowledge on the potential causes of this variability, we determined 18O-kinetic isotope effects (KIEs) across a broad range of O2-consuming enzymes. The studied enzymes included nine flavin-dependent, five copper-dependent, and one copper-heme-dependent oxidases, as well as one flavin-dependent monooxygenase. For twelve of these enzymes, 18O-KIEs were determined for the first time. The comparison of 18O-KIEs, determined in this and previous studies, to calculated 18O-equilibrium isotope effects revealed distinct patterns of O-isotopic fractionation within and between enzyme groups, reflecting differences in active-site structures and O2-reduction mechanisms. Flavin-dependent O2-consuming enzymes exhibited two distinct ranges of 18O-KIEs (from 1.020 to 1.034 and from 1.046 to 1.058), likely associated with the rate-limiting steps of two different O2-reduction mechanisms (sequential vs. concomitant 2-electron transfer). In comparison, iron- and copper-dependent enzymes displayed a narrower range of 18O-KIEs, with overall lower values (from 1.009 to 1.028), which increased with the degree of O2 reduction during the rate-limiting step. Similar to flavin-dependent O2-consuming enzymes, copper-dependent O2-consuming enzymes also featured two main, yet narrower, ranges of 18O-KIEs (from 1.009 to 1.010 and from 1.017 to 1.022), likely associated with the rate-limiting formation of a copper-superoxo or copper-hydroperoxo intermediate. Overall, our findings support generalizations regarding expected 18O-KIEs ranges imparted by O2-consuming enzymes and have the potential to help interpret stable O2 isotopic fractionation patterns across different environmental scales.

Publisher's note: Copernicus Publications remains neutral with regard to jurisdictional claims made in the text, published maps, institutional affiliations, or any other geographical representation in this paper. While Copernicus Publications makes every effort to include appropriate place names, the final responsibility lies with the authors. Views expressed in the text are those of the authors and do not necessarily reflect the views of the publisher.
Share

Journal article(s) based on this preprint

12 Sep 2025
Variability in oxygen isotopic fractionation of enzymatic O2 consumption
Carolina F. M. de Carvalho, Moritz F. Lehmann, and Sarah G. Pati
Biogeosciences, 22, 4579–4600, https://doi.org/10.5194/bg-22-4579-2025,https://doi.org/10.5194/bg-22-4579-2025, 2025
Short summary
Download

The requested preprint has a corresponding peer-reviewed final revised paper. You are encouraged to refer to the final revised version.

Short summary
Using O2 stable isotope analysis, we determined the isotopic fractionation of biological O2...
Share