Impact of metabolism and temperature on 2H/1H fractionation in lipids of marine bacterium Shewanella piezotolerans WP3

Chen, Xin; Zhao, Weishu; Dong, Liang; Jian, Huahua; Liang, Lewen; Wang, Jing; Wang, Fengping

doi:https://doi.org/10.5194/bg-2022-232

Preprints

https://doi.org/10.5194/bg-2022-232

Preprints

12 Dec 2022

Status: this preprint is currently under review for the journal BG.

Impact of metabolism and temperature on ²H/¹H fractionation in lipids of marine bacterium Shewanella piezotolerans WP3

Xin Chen¹, Weishu Zhao², Liang Dong¹, Huahua Jian², Lewen Liang², Jing Wang¹, and Fengping Wang^1,2,3 Xin Chen et al.

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¹School of Oceanography, Shanghai Jiao Tong University, Shanghai,200240, China
²State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China
³Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Zhuhai, 519000, China

Received: 05 Dec 2022 – Discussion started: 12 Dec 2022

Abstract. Compound-specific hydrogen isotopes are increasingly used as a powerful proxy for investigating biogeochemical cycle and climate change over the past two decades. Understanding the hydrogen isotope in extant organisms is fundamental for us to interpret such isotope signals preserved in natural environmental samples. Here, we studied the controls on hydrogen isotope fractionation between fatty acids and growth water by a Fe-reducing heterotrophic marine bacterium Shewanella piezotolerans WP3 growing on different organic substrates, including N-acetyl-D-glucosamine (GlcNac), glucose, acetate, pyruvate, L-alanine and L-glutamate. Meanwhile, we also evaluated the impact of growth temperature on the hydrogen isotope composition of fatty acids using GlcNac as sole organic substrate. Our results show that the abundance-weighted mean fatty acids/water fractionations (ε_FA/Water) display considerable variations for cultures grown on different substrates. Specifically, WP3 yielded the most ²H-enriched fatty acids growing on L-glutamate and pyruvate with ε_FA/Water of 52 ± 14 ‰ and 44 ± 4 ‰ respectively, and exhibited ²H-depleted using GlcNac (-76 ± 1 ‰) and glucose (-67 ± 35 ‰) as sole carbon sources, relatively small fractionations on acetate (23 ± 3 ‰) and L-alanine (-4 ± 9 ‰). Combined with metabolic model analysis, our results indicate that the central metabolic pathways exert a fundamental effect on the hydrogen isotope composition of fatty acids in heterotrophs. Temperature also has obvious influence on the δ²H values of fatty acids, with strongly ²H-depleted at optimal growth temperature (15 and 20 °C) and relatively small fractionations at non-optimal temperatures (4, 10, and 25 °C). We hypothesized that it is most likely controlled by the temperature effects on the activity of associated enzymes for NADPH production. This study helps understanding the controlling factors of hydrogen isotope fractionation by marine bacteria, lays the foundation for further interpreting the hydrogen isotope signatures of lipids as an important proxy to decode the biogeochemical cycles and ecological changes in marine sediments.

Xin Chen et al.

Status: final response (author comments only)

RC1: 'Comment on bg-2022-232', Anonymous Referee #1, 03 Jan 2023

I have carefully read the paper entitled ”” by Chen et al. The authors studied the controls on hydrogen isotope fractionation between fatty acids and frowth water by a Fe-reducing heterotrophic marine bacterium WP3 The authors also evaluated the impact of growth temperature on hydrogen isotope fractionations. Given the potential of hydrogen isotopes as a proxy in the geological records, more studies on factors controlling hydrogen fractionation bacteria are needed. The cultures are reasonable, the data are sound and suitable for publication on Biogeosciences after minor revisions. My mainly concern is that the biomarker compositions are different when supplied with different substrates or temperatures, the authors need to address the reasons and if those reasons influence the hydrogen isotope fractionation as well. Although the manuscript is overall well written, the language is needed to be polished further.

Minor errors as follow:

Line 172, branched chain lengths can be saturated fatty acids, too. If you mean to highlight branched chain lengths, you need to say “compared to straight chain lengths”. The authors compared the hydrogen isotopes of iC15 and aiC15 with C16 and conclude different precursors make different hydrogen fractionations, the authors need to clarify who are their precursors and what pathways they used to produce these fatty acids.

Since the δ²H values are closely related to the NADPH to NADH, is it possible to determine the δ²H values of NADPH to NADH? Especially supply different bacteria with the same substrates.

Line 101, ……described in Rodríguez-Ruiz et al. (1998).

Line 281, relatively larger

Citation: https://doi.org/10.5194/bg-2022-232-RC1
RC2: 'Comment on bg-2022-232', Anonymous Referee #2, 07 Jan 2023

In this paper, Chen et al., performed a culture experiment on heterotrophic marine bacterium WP3 that widely occurs in the deep sea, with different organic substates and a temperature gradient, to study the impact of metabolism and temperature on the microbial lipid biomarker (-fatty acids) hydrogen isotope fractionation. They showed that central metabolic pathways associated with NADPH production exert an important effect in determining the hydrogen isotope fractionation, and temperature play a secondary role. This study is very valuable for understanding compound-specific hydrogen isotopes in marine sediment records for biogeochemical and paleoclimate studies. I am supportive of publication after consideration of following minor comments.

Minor comments

L20: Add “and” before “relatively small”.

L23-24: Specifically add how much hydrogen isotope fractionations are observed under different temperatures.

L25: Please rephrase this sentence “it is most likely controlled……”. For example, “We hypothesized that this may associated with temperature-induced enzyme activity……”.

L33: Add ‘are assumed” after “phototrophic algae”.

L34-37: Please revise this sentence. For example, “However, increasing studies have found that there are large ranges of hydrogen isotope ratios in lipids (up to 700‰) from variation environmental samples,……”. Note that the fractionation values are up to 700‰, rather than 700%.

L41: Change “thrived” to “thriving”.

L74-76: Dose belongs to Shewanella? If so, please specifically address this here.

L89: Please add some sentences: what and where marine samples does S. piezotolerans WP3 to be isolated and enriched?

L104: How did you derivatize fatty acids into FAMEs? Please add the chemical experimental process.

L110: How much temperature did you use in the pyrolysis interface?

L115: Add the correction formula and hydrogen isotope value of methyl group.

L130-140: Add some statistical box chart figures as supplementary materials to show the differences in hydrogen isotope fractionation of fatty acids under different substrates.

L225: Change “grown” to “growing”.

L265-267: Different individual fatty acids have substantial differences in hydrogen isotope fractionation. Please add some figures as supplementary materials to show the changes in hydrogen isotope fractionation of same individual fatty acid along the temperature gradient.

L273-274: Please rephrase this sentence. For example, “The mechanisms may associated with growth rates and enzyme activities in organisms controlled by temperature.”

L324-325: Please revise the grammar.

L326: Change “growing on sugars” to “using sugars as substrates”.

L381: Change “growing on” to “using”.

Citation: https://doi.org/10.5194/bg-2022-232-RC2

Xin Chen et al.

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Short summary

Here, we studied the effects of metabolism and growth temperature on ²H/¹H fractionation between fatty acids and growth water (ε_FA/Water) by Shewanella piezotolerans WP3. Our results show that the ε_FA/Water values display considerable variations for cultures grown on different substrates. Combined with metabolic model analysis, our results indicate that the central metabolic pathways exert a fundamental effect on the hydrogen isotope composition of lipids in heterotrophs.


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