ArticlePloS one2013
Using synthetic biology to distinguish and overcome regulatory and functional barriers related to nitrogen fixation.
Article in PloS one, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 25 papers.
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Who cites it
25 citing papers in PubMed, 50 citations in OpenAlex.
- Use of translational fusions to express functional Klebsiella oxytoca dinitrogenase reductase in plant mitochondria.Plant molecular biology · 2026Article
- Predicting compatibility between ferredoxins and the Fe protein of nitrogenase using in silico protein modeling.Protein science : a publication of the Protein Society · 2026Article
- Article
- Organelle-dependent polyprotein designs enable stoichiometric expression of nitrogen fixation components targeted to mitochondria.Proceedings of the National Academy of Sciences of the United States of America · 2023Article
- Engineering Nitrogenases for Synthetic Nitrogen Fixation: From Pathway Engineering to Directed Evolution.Biodesign research · 2023Review
- Article
- A colorimetric method to measure in vitro nitrogenase functionality for engineering nitrogen fixation.Scientific reports · 2022Article
- The Chloroplast ofInternational journal of molecular sciences · 2021Review
- Nitrogen fixation in maize: breeding opportunities.TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik · 2021Review
- Minimal gene set fromProceedings of the National Academy of Sciences of the United States of America · 2021Article
- Exploiting Biological Nitrogen Fixation: A Route Towards a Sustainable Agriculture.Plants (Basel, Switzerland) · 2020Review
- Biological nitrogen fixation in maize: optimizing nitrogenase expression in a root-associated diazotroph.Journal of experimental botany · 2020Article
- Using synthetic biology to overcome barriers to stable expression of nitrogenase in eukaryotic organelles.Proceedings of the National Academy of Sciences of the United States of America · 2020Article
- Engineered CRISPRa enables programmable eukaryote-like gene activation in bacteria.Nature communications · 2019Article
- Extreme bioengineering to meet the nitrogen challenge.Proceedings of the National Academy of Sciences of the United States of America · 2018Article
- Polyprotein strategy for stoichiometric assembly of nitrogen fixation components for synthetic biology.Proceedings of the National Academy of Sciences of the United States of America · 2018Article
- Global investigation of an engineered nitrogen-fixing Escherichia coli strain reveals regulatory coupling between host and heterologous nitrogen-fixation genes.Scientific reports · 2018Article
- State of the art in eukaryotic nitrogenase engineering.FEMS microbiology letters · 2018Review
- Formation of Nitrogenase NifDK Tetramers in the Mitochondria of Saccharomyces cerevisiae.ACS synthetic biology · 2017Article
- Modular electron-transport chains from eukaryotic organelles function to support nitrogenase activity.Proceedings of the National Academy of Sciences of the United States of America · 2017Article
Corrections and comments
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Authors and funding
7 authors at 3 institutions in 2 countries.
Funding
Abstract
Biological nitrogen fixation is a complex process requiring multiple genes working in concert. To date, the Klebsiella pneumoniae nif gene cluster, divided into seven operons, is one of the most studied systems. Its nitrogen fixation capacity is subject to complex cascade regulation and physiological limitations. In this report, the entire K. pneumoniae nif gene cluster was reassembled as operon-based BioBrick parts in Escherichia coli. It provided ~100% activity of native K. pneumoniae system. Based on the expression levels of these BioBrick parts, a T7 RNA polymerase-LacI expression system was used to replace the σ(54)-dependent promoters located upstream of nif operons. Expression patterns of nif operons were critical for the maximum activity of the recombinant system. By mimicking these expression levels with variable-strength T7-dependent promoters, ~42% of the nitrogenase activity of the σ(54)-dependent nif system was achieved in E. coli. When the newly constructed T7-dependent nif system was challenged with different genetic and physiological conditions, it bypassed the original complex regulatory circuits, with minor physiological limitations. Therefore, we have successfully replaced the nif regulatory elements with a simple expression system that may provide the first step for further research of introducing nif genes into eukaryotic organelles, which has considerable potentials in agro-biotechnology.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.