Exploiting genetic diversity and gene synthesis to identify superior nitrogenase NifH protein variants to engineer N2-fixation in plants

• DOI: 10.1038/s42003-020-01536-6
• Publication Date: 2021-01-04T11:02:41Z
• Authors: Xi Jiang; Lucía Payá-Tormo; Diana Coroian; Inés García-Rubio; Rocío Castellanos...; Álvaro Eseverri; Gema López-Torrejón; Stefan Burén; Luis Manuel Rubio
• Keywords: Nicotiana; 2. Zero hunger; 0303 health sciences; Bacteria; QH301-705.5; Biología; Iron; Saccharomyces cerevisiae; http://metadata.un.org/sdg/3; Article; Mitochondria; 03 medical and health sciences; Nitrogen Fixation; Genetic engineering; Tobacco; Nitrogenase; Biology (General); Genetic Engineering; Ensure healthy lives and promote well-being for all at all ages; Gene Library

Abstract Engineering nitrogen fixation in eukaryotes requires high expression of functional nitrogenase structural proteins, a goal that has not yet been achieved. Here we build knowledge-based library containing 32 nifH sequences from prokaryotes diverse ecological niches and metabolic features combine with rapid screening tobacco to identify superior NifH variants for plant mitochondria expression. Three outperform are further tested yeast. Hydrogenobacter thermophilus (Aquificae) is isolated large quantities yeast fulfills protein requirements efficient N 2 fixation, including electron transfer substrate reduction, P-cluster maturation, FeMo-co biosynthesis. H. expressed leaves shows lower activity than However, [Fe 4 S ] clusters NifU vitro increases 10-fold the tobacco-isolated NifH, revealing [Fe-S] cluster availability constitutes bottleneck engineer nitrogenases.