Browsing by Author "Crumbley, Anna Morgan"

Now showing 1 - 1 of 1
  • Thumbnail Image
    Item
    Methane Bioreforming for the Biosynthesis of Reduced Molecules at Ambient Pressure and Temperature
    (2020-07-30) Crumbley, Anna Morgan; Gonzalez, Ramon; San, Ka-Yiu; Segatori, Laura
    The inherent principles of biology offer access to low temperature and pressure reactions with high selectivity in single-reactor vessels as an efficient route to chemical production through value-added biomanufacturing. Remote methane (CH4), a low-cost, but high-energy, gaseous hydrocarbon comprises a potentially-attractive feedstock distributed globally and produced by both renewable and traditional sources. Utilizing CH4 as a feedstock for value-added biomanufacturing of products such as ammonia (NH3), an expensive but essential source of N in agriculture, would facilitate NH3 production in remote areas by addressing current technological challenges related to high temperature and pressure systems. This work details the development of a biological pathway producing NH3 from CH4, oxygen (O2), and nitrogen (N2) in the air at ambient pressure and temperature. A proof-of-concept system discussed herein demonstrates a syntrophic microbial consortium of Azotobacter vinelandii M5I3 and Methylomicrobium buryatense 5GB1 pAMR4-dtom1 performing methane bioreforming (MBR) and powering biological nitrogen fixation (BNF) through extracellular carbon and energy transfer. Using a co-culture of bacteria synthetically engineered to secrete extracellular electron carrier lactic acid and NH3 derived from CH4 and air, this work demonstrates accumulated extracellular carbon and nitrogen products in the proof-of-concept system. Through a combination of iterative refinement, control, and stable-isotope 15N2 labeling experiments, the work further demonstrates active BNF in the co-culture, albeit at a low level. The downstream portion of the pathway was optimized using in silico kinetic modeling tools and experimental synthetic biology modifications to enhance NH3 production by the system further. While overall NH3 yields remain low after implementing these modifications, in part due to gas-solubility mass transfer challenges, this work suggests that carbon and energy present in CH4 are capable of being utilized in a methane bioreforming manner to power enzymatic activity beyond carbon-based product generation. Furthermore, electron transfer to nitrogenase represents a bottleneck for efficient NH3 production in native biological nitrogen-fixing organisms.