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Minimal Influence of [NiFe] Hydrogenase on Hydrogen Isotope Fractionation in H_2-Oxidizing Cupriavidus necator

Campbell, Brian J. and Sessions, Alex L. and Fox, Daniel N. and Paul, Blair G. and Qin, Qianhui and Kellermann, Matthias Y. and Valentine, David L. (2017) Minimal Influence of [NiFe] Hydrogenase on Hydrogen Isotope Fractionation in H_2-Oxidizing Cupriavidus necator. Frontiers in Microbiology, 8 . Art. No. 1886. ISSN 1664-302X. PMCID 29085342.

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Fatty acids produced by H_2-metabolizing bacteria are sometimes observed to be more D-depleted than those of photoautotrophic organisms, a trait that has been suggested as diagnostic for chemoautotrophic bacteria. The biochemical reasons for such a depletion are not known, but are often assumed to involve the strong D-depletion of H_2. Here, we cultivated the bacterium Cupriavidus necator H16 (formerly Ralstonia eutropha H16) under aerobic, H_2-consuming, chemoautotrophic conditions and measured the isotopic compositions of its fatty acids. In parallel with the wild type, two mutants of this strain, each lacking one of two key hydrogenase enzymes, were also grown and measured. In all three strains, fractionations between fatty acids and water ranged from -173‰ to -235‰, and averaged -217‰, -196‰, and -226‰, respectively, for the wild type, SH- mutant, and MBH- mutant. There was a modest increase in δD as a result of loss of the soluble hydrogenase enzyme. Fractionation curves for all three strains were constructed by growing parallel cultures in waters with δD_(water) values of approximately -25‰, 520‰, and 1100‰. These curves indicate that at least 90% of the hydrogen in fatty acids is derived from water, not H_2. Published details of the biochemistry of the soluble and membrane-bound hydrogenases confirm that these enzymes transfer electrons rather than intact hydride (H-) ions, providing no direct mechanism to connect the isotopic composition of H_2 to that of lipids. Multiple lines of evidence thus agree that in this organism, and presumably others like it, environmental H_2 plays little or no direct role in controlling lipid δD values. The observed fractionations must instead result from isotope effects in the reduction of NAD(P)H by reductases with flavin prosthetic groups, which transfer two electrons and acquire H+ (or D+) from solution. Parallels to NADPH reduction in photosynthesis may explain why D/H fractionations in C. necator are nearly identical to those in many photoautotrophic algae and bacteria. We conclude that strong D-depletion is not a diagnostic feature of chemoautotrophy.

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Sessions, Alex L.0000-0001-6120-2763
Additional Information:© 2017 Campbell, Sessions, Fox, Paul, Qin, Kellermann and Valentine. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Received: 06 October 2016; Accepted: 14 September 2017; Published: 04 October 2017. Author Contributions: BC planned the experiments, conducted research, analyzed the data, and wrote the manuscript. AS performed lipid analyses, analyzed the data, and wrote the manuscript. DF helped with microbial cultures and gas isotope analyses. BP provided analysis of genomic data for strain H16. QQ cultivated and sampled microbial cultures. MK performed the polar lipid analysis, DV planned and supervised the experiments, analyzed the data, and edited the manuscript. This work was supported by the NASA Graduate Student Researcher Program (to BC) and grants from the National Science Foundation (EAR-0311894 to DV and AS; OCE-1046144 to DV; EAR-1529120 to AS). Conflict of Interest Statement: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors gratefully acknowledge Robert Petty, Chao Li, and Fenfang Wu for assistance with various isotopic analyses. We thank the three reviewers, who contributed substantively to improving the manuscript.
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NASA Graduate Student Research FellowshipUNSPECIFIED
Subject Keywords:Cupriavidus necator, hydrogenase, hydrogen isotope, D/H, fatty acid, autotrophic metabolism
PubMed Central ID:29085342
Record Number:CaltechAUTHORS:20171020-085602582
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Official Citation:Campbell BJ, Sessions AL, Fox DN, Paul BG, Qin Q, Kellermann MY and Valentine DL (2017) Minimal Influence of [NiFe] Hydrogenase on Hydrogen Isotope Fractionation in H2-Oxidizing Cupriavidus necator. Front. Microbiol. 8:1886. doi: 10.3389/fmicb.2017.01886
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:82535
Deposited By: Tony Diaz
Deposited On:20 Oct 2017 17:20
Last Modified:03 Oct 2019 18:55

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