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Exploiting gene diversity to identify superior nitrogenase reductase variants to engineer nitrogen fixing plants

Plants absorb nitrogen from the soil or through symbiotic relationships with nitrogen fixing bacteria. Obtaining nitrogen fixing crop varieties would reduce the use of nitrogen based fertilizers drastically and therefore, reducing the negative effects associated to the use of these fertilizers in the environment while maintaining the crop yields high. This objective could be achieved by transferring the bacterial genes coding the nitrogenase enzyme to plants. Protein insolubility was the main problem detected when expressing the NifH component of nitrogenase of the model organism Azotobacter vinelandii in tobacco mitochondria.

In this work 32 nifH gene variants from 32 different organisms and diverse ecological niches were synthesized. Solubility of these variants were analyzed when expressed in tobacco mitochondria and 3 of them were outstanding in terms of protein solubility. In order to analyze the biochemical properties of the 3 candidates, they were expressed in yeast, a simpler model organism than plants. Hydrogenobacter thermophilus NifH variant was the candidate that demonstrated to have the biggest potential to be a functional active enzyme. This protein variant was purified from tobacco mitochondria and protein activity could be detected when the purification was performed after the end of the dark period and addition of iron fertilizer to the soil.

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Original Paper:

Jiang, X., Payá-Tormo, L., Coroian, D., García-Rubio, I., Castellanos-Rueda, R., Eseverri, Á., López-Torrejón, G., Burén, S., Rubio, L.M. 2021. Exploiting genetic diversity and gene synthesis to identify superior nitrogenase NifH protein variants to engineer N 2 -fixation in plants. Communications Biology 4, 1–11. DOI: 10.1038/s42003-020-01536-6

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