Engineered Geobacter sulfurreducens Achieves a Record-High Rate of Biological Nitrogen Fixation

Biological nitrogen fixation (BNF) represents a sustainable alternative to the capital- and energy-intensive Haber-Bosch process, which dominates industrial fertilizer production but contributes significantly to global carbon emissions. While Geobacter sulfurreducens has been identified as a promising diazotroph, its BNF efficiency remains untapped. Here, we engineered G. sulfurreducens by deleting the ammonium transporter gene amtB (strain ΔamtB), achieving a record-high BNF rate of 20.57 ± 0.87 mg N L-1 day-1, approximately twice that of the wild-type strain and exceeding those of all reported free-living diazotrophs and synthetic catalysts for ambient nitrogen fixation. 15N isotope labeling and nitrogenase activity assays confirmed enhanced fixation, whereas biofertilizer trials demonstrated the ability of ΔamtB as a biofertilizer to support Arabidopsis thaliana growth. Mechanistic studies revealed that amtB deletion (1) amplified cellular nitrogen assimilation, (2) alleviated ammonium-mediated suppression of nitrogenase activity, and (3) disrupted energy-intensive futile ammonium cycling, conserving cell energy. This study establishes ΔamtB as a scalable BNF platform for sustainable agriculture, offering a viable pathway to decarbonize fertilizer production.