For optimal yields, crop plants require fixed nitrogen in the form of ammonia or nitrate fertilizers, but this requires large fossil fuel inputs and can also result in runoff which contaminates aquifers and estuaries. Unlike some microbes that have the capacity to fix atmospheric nitrogen, plants do not have this ability. The ultimate goal of this research is to engineer a novel synthetic nitrogen fixing organelle, with the long-term aim of conferring efficient nitrogen fixation in non-leguminous crop plants. However, there are significant hurdles before realizing this goal, which include high metabolic energy costs and overcoming oxygen sensitivity of the process. In this project, tools of synthetic biology will be used to engineer nitrogen fixation into a simple model system. Cyanobacteria are single-celled organisms that are evolutionarily related to plant plastids. In cyanobacteria, the engineering goals should be tractable, constituting a technological stepping stone that would lead to the engineering of nitrogen fixation into plant plastids. For this project to be successful, several objectives need to be met. First, ideal candidate gene clusters required for nitrogen fixation need to be identified. Second, using this information, tunable nitrogen-fixing gene modules, which can be precisely controlled, will be built and moved into cyanobacteria. Finally, to deal with the high metabolic energy costs of the process, a novel strategy will be employed by which extra light absorption capacity is engineered into cyanobacteria. These objectives are complex and multi-faceted, requiring tight coordination between participating laboratories. Successful completion of this research will lead to an engineered synthetic, controllable nitrogen fixing gene cluster linked energetically to light energy, which can ultimately be transferred into plastids of crop plants in the form of a 'nitroplast'.
BROADER IMPACTS One of the major challenges of the twenty-first century is to ensure food security for the world's people. At the core of this challenge is the problem of nitrogen assimilation by non-leguminous crop plants. The goal of this project is to build a novel synthetic, controllable nitrogen-fixing module into a cyanobacterium. To achieve this goal, a team of scientific experts have been assembled from the fields of biophysics, biochemistry, molecular genetics and synthetic biology. The outcome of this research is expected to benefit basic researchers in academia as well as applied scientists through the development of new tools for cyanobacteria, and through products such as the introduction of nitrogen fixation into plants. This project is a unique opportunity for methodology exchange between U.S and U.K. scientists and for developing tools that will be freely available to the synthetic biology and cyanobacterial research communities. If successful, the work will also benefit traditional agricultural research. The preparation of the next generation of scientists is a major goal of this interdisciplinary team. The research will benefit higher education through intensive research training at the undergraduate, graduate and postdoctoral levels and through international exchange between US and UK collaborators, publications and presentations at scientific meetings. The new technologies derived from this work should provide tools and knowledge to boost nitrogen fixation capacity that could strongly impact agriculture.