Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity
Abstract Biological nitrogen fixation (BNF) by canonical molybdenum and complementary vanadium and iron-only nitrogenase isoforms is the primary natural source of newly fixed nitrogen. Understanding controls on global nitrogen cycling requires knowledge of the isoform responsible for environmental B...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Nature Portfolio
2022-12-01
|
| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-022-24860-9 |
| _version_ | 1797977480845328384 |
|---|---|
| author | Shannon J. Haynes Romain Darnajoux Eunah Han Sergey Oleynik Ezra Zimble Xinning Zhang |
| author_facet | Shannon J. Haynes Romain Darnajoux Eunah Han Sergey Oleynik Ezra Zimble Xinning Zhang |
| author_sort | Shannon J. Haynes |
| collection | DOAJ |
| description | Abstract Biological nitrogen fixation (BNF) by canonical molybdenum and complementary vanadium and iron-only nitrogenase isoforms is the primary natural source of newly fixed nitrogen. Understanding controls on global nitrogen cycling requires knowledge of the isoform responsible for environmental BNF. The isotopic acetylene reduction assay (ISARA), which measures carbon stable isotope (13C/12C) fractionation between ethylene and acetylene in acetylene reduction assays, is one of the few methods that can quantify isoform-specific BNF fluxes. Application of classical ISARA has been challenging because environmental BNF activity is often too low to generate sufficient ethylene for isotopic analyses. Here we describe a high sensitivity method to measure ethylene δ13C by in-line coupling of ethylene preconcentration to gas chromatography-combustion-isotope ratio mass spectrometry (EPCon-GC-C-IRMS). Ethylene requirements in samples with 10% v/v acetylene are reduced from > 500 to ~ 20 ppmv (~ 2 ppmv with prior offline acetylene removal). To increase robustness by reducing calibration error, single nitrogenase-isoform Azotobacter vinelandii mutants and environmental sample assays rely on a common acetylene source for ethylene production. Application of the Low BNF activity ISARA (LISARA) method to low nitrogen-fixing activity soils, leaf litter, decayed wood, cryptogams, and termites indicates complementary BNF in most sample types, calling for additional studies of isoform-specific BNF. |
| first_indexed | 2024-04-11T05:08:39Z |
| format | Article |
| id | doaj.art-8421511beb6847f6b8d169851cc118f0 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-04-11T05:08:39Z |
| publishDate | 2022-12-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-8421511beb6847f6b8d169851cc118f02022-12-25T12:11:52ZengNature PortfolioScientific Reports2045-23222022-12-0112111210.1038/s41598-022-24860-9Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activityShannon J. Haynes0Romain Darnajoux1Eunah Han2Sergey Oleynik3Ezra Zimble4Xinning Zhang5Department of Geosciences, Princeton UniversityDepartment of Geosciences, Princeton UniversityDepartment of Geosciences, Princeton UniversityDepartment of Geosciences, Princeton UniversityHigh Meadow Environmental Institute, Princeton UniversityDepartment of Geosciences, Princeton UniversityAbstract Biological nitrogen fixation (BNF) by canonical molybdenum and complementary vanadium and iron-only nitrogenase isoforms is the primary natural source of newly fixed nitrogen. Understanding controls on global nitrogen cycling requires knowledge of the isoform responsible for environmental BNF. The isotopic acetylene reduction assay (ISARA), which measures carbon stable isotope (13C/12C) fractionation between ethylene and acetylene in acetylene reduction assays, is one of the few methods that can quantify isoform-specific BNF fluxes. Application of classical ISARA has been challenging because environmental BNF activity is often too low to generate sufficient ethylene for isotopic analyses. Here we describe a high sensitivity method to measure ethylene δ13C by in-line coupling of ethylene preconcentration to gas chromatography-combustion-isotope ratio mass spectrometry (EPCon-GC-C-IRMS). Ethylene requirements in samples with 10% v/v acetylene are reduced from > 500 to ~ 20 ppmv (~ 2 ppmv with prior offline acetylene removal). To increase robustness by reducing calibration error, single nitrogenase-isoform Azotobacter vinelandii mutants and environmental sample assays rely on a common acetylene source for ethylene production. Application of the Low BNF activity ISARA (LISARA) method to low nitrogen-fixing activity soils, leaf litter, decayed wood, cryptogams, and termites indicates complementary BNF in most sample types, calling for additional studies of isoform-specific BNF.https://doi.org/10.1038/s41598-022-24860-9 |
| spellingShingle | Shannon J. Haynes Romain Darnajoux Eunah Han Sergey Oleynik Ezra Zimble Xinning Zhang Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity Scientific Reports |
| title | Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity |
| title_full | Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity |
| title_fullStr | Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity |
| title_full_unstemmed | Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity |
| title_short | Quantification of biological nitrogen fixation by Mo-independent complementary nitrogenases in environmental samples with low nitrogen fixation activity |
| title_sort | quantification of biological nitrogen fixation by mo independent complementary nitrogenases in environmental samples with low nitrogen fixation activity |
| url | https://doi.org/10.1038/s41598-022-24860-9 |
| work_keys_str_mv | AT shannonjhaynes quantificationofbiologicalnitrogenfixationbymoindependentcomplementarynitrogenasesinenvironmentalsampleswithlownitrogenfixationactivity AT romaindarnajoux quantificationofbiologicalnitrogenfixationbymoindependentcomplementarynitrogenasesinenvironmentalsampleswithlownitrogenfixationactivity AT eunahhan quantificationofbiologicalnitrogenfixationbymoindependentcomplementarynitrogenasesinenvironmentalsampleswithlownitrogenfixationactivity AT sergeyoleynik quantificationofbiologicalnitrogenfixationbymoindependentcomplementarynitrogenasesinenvironmentalsampleswithlownitrogenfixationactivity AT ezrazimble quantificationofbiologicalnitrogenfixationbymoindependentcomplementarynitrogenasesinenvironmentalsampleswithlownitrogenfixationactivity AT xinningzhang quantificationofbiologicalnitrogenfixationbymoindependentcomplementarynitrogenasesinenvironmentalsampleswithlownitrogenfixationactivity |