CAREER:The Role of ATP Hydrolysis in Biological Nitrogen Fixation

University Of California-San Diego, La Jolla CA

Investigators

Abstract

Reduced forms of molecular nitrogen (dinitrogen) are essential for the production of fertilizers and countless industrial chemicals, as well as the biosynthesis of amino and nucleic acids. The reduction of dinitrogen to ammonia (nitrogen fixation) by the Haber-Bosch process requires high temperatures and pressures, consuming over 100 billion watts of power every year. Biological dinitrogen fixation does not require such extreme conditions; indeed, the enzyme nitrogenase catalyzes ammonia synthesis under ambient conditions. After decades of intense research efforts, however, it is still not known how nitrogenase activates dinitrogen. Nitrogenase catalysis is distinct from other multielectron/multiproton catalytic reactions in its requirement of 16 ATP molecules per turnover reaction despite a favorable thermodynamic driving force. This project aims to elucidate why and how ATP hydrolysis is required in biological nitrogen fixation. Recent structural studies show that the two constituents of nitrogenase, the Fe-protein (electron donor/ATPase) and the MoFe-protein (catalytic component), can assume at least three docking geometries, depending on the ATP-hydrolysis state. By developing and utilizing several powerful chemical and biophysical tools, this project will probe whether multiple Fe-protein:MoFe-protein docking modes are functionally important, and if they are involved in timing the successive electron and proton transfers into the catalytic metal cluster (FeMoco). In parallel, photochemical methods will be utilized to investigate the possibility of driving substrate reduction at FeMoco without requiring ATP hydrolysis. The demonstration of photoactivation of nitrogenase will open new avenues for studying its mechanism that in turn could lead to the development of biocatalytic systems for ammonia and hydrogen production. Broader Impacts: The complexity of biological nitrogen fixation requires a multidisciplinary plan of attack. This project combines a multitude of experimental approaches that will provide an expansive training ground for graduate and undergraduate students. Biological nitrogen fixation sustains a large fraction (40%) of the world's population, and the industrial Haber-Bosch process is responsible for considerable amounts of energy consumption and greenhouse gas emissions. A thorough understanding of nitrogenase mechanism could lead to the design of clean and efficient biocatalysts for ammonia production, which would have an immense economic and environmental impact. Nitrogen fixation also provides a conduit into the education goals of this project, which is to raise the awareness of students about the global energy problem, and to train scientists in energy biosciences. These goals will be addressed on several fronts, including a) the interdisciplinary training of graduate and undergraduate students in the laboratory, b) restructuring of an advanced course in Bioinorganic Chemistry to focus on redox-catalytic processes involved in global carbon, nitrogen, oxygen, and sulfur cycles, as well as the design of a multidisciplinary course on Global Energy Problem and Alternative Energy Research, and c) outreach to low-income students from underrepresented groups attending a local charter school, and their recruitment through seminars.