Photo-assembly and Efficiency of Photosynthetic Water Oxidases: Probing the Catalytic Core Atom by Atom

In this award from the Chemistry of Life Processes Program in the Chemistry Division, Drs. G. Charles Dismukes and Gennady M. Ananyev, from Rutgers, The State University of New Jersey, seek to understand the physico-chemical principles that govern the assembly of the inorganic cofactors to form the water-oxidation complex (WOC) within Photosystem II (PSII) of photosynthetic organisms. This complex is nature's sole enzymatic solution for splitting water into molecular oxygen, protons, and electrons using sunlight. Despite apparent conservation of the WOC catalytic core, comprised of a Mn4CaO5 cluster, two chloride ions, and proximal amino acid residues, large differences in catalytic turnover rates occur between PSII species in vivo, indicative of functional differences that are not yet understood. The general experimental approach of this project uses organismal, biochemical and inorganic chemical strategies. The expected outcome will be to determine the allowed range of possible non-native inorganic cofactors that can support catalytic functioning of PSII-WOCs, determine their relative binding affinities and sites, and measure their kinetic performance in catalysis across different species. A long term goal is to understand the chemical basis for the evolutionary conservation of this enzyme class. These studies will be conducted using model photosynthetic organisms and using both native cells (in vivo) and isolated PSII complexes (in vitro). A number of complementary techniques will be used that allow comparison of kinetic performance over a wide range of flashing frequencies including: ultra-sensitive electrochemical O2 concentration and rate measurements, chlorophyll-detected Fast Repetition Rate fluorometry, and dye-detected kinetic pH measurements. Because no single inorganic component of the WOC works independently, we will conduct a systematic 'atom-by-atom' analysis that reveals their synergistic contributions.

Advances in our understanding of the mechanism of photosynthetic water oxidation will inform alternative energy research in two areas. First, insights from the experiments proposed here may guide the design of genetically modified photosynthetic organisms that, for example, are expected to have improved biomass accumulation. Second, elucidation of the principles of photosynthetic water oxidation will guide the design of water oxidation catalysts for artificial photosynthesis systems capable of producing renewable fuels as alternative to fossil fuels. The proposed studies would allow research training of Rutgers undergraduate and postgraduate students, including women and those of underrepresented backgrounds, for future careers and advanced training in the renewable energy field. These proposed studies will leverage educational training programs sponsored by two NSF IGERT grants at Rutgers. The new instruments built and upgrades to existing instruments described in this proposal will improve the infrastructure available in our laboratory and accommodate other users at Rutgers, while supporting multiple collaborators and their funding agencies.