The Interaction of Calcium and Chloride at the Oxygen Evolving Complex of Photosystem II

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Intellectual MeritIn the utilization of energy on earth, photosynthesis by higher plants has a profound effect. It is the source of hydrocarbon energy for nearly all life forms through the capture of the sun's light energy and it is almost the sole source of dioxygen for respiration. A thorough understanding of the photosynthetic process is central to our ability to provide for our own energy needs in the future. Photosystem II is one of the two major light absorbing reaction centers of higher plants and cyanobacteria. It also contains the site of oxygen evolution, an unusual Mn4Ca catalytic center. The mechanism of the multiple step conversion of water into dioxygen is still not well understood. Both calcium and chloride are important participants in this process and they appear to interact with each other during oxygen production. As a part of the Mn4Ca cluster, calcium probably has a direct role in oxygen formation. Although chloride is not directly coordinated to the Mn4Ca cluster, it is required for transitions to higher oxidation states and appears to stabilize the calcium ion. This project is intended to help clarify the role of calcium in photosystem II of higher plants and its relationship to activation by chloride. The approach used will be to characterize the effects of calcium depletion or disruption at the Mn4Ca catalytic center. Two methods for biochemically disturbing the function of calcium will be used: treatment with mildly reduced pH and moderately high sodium chloride concentration; and treatment with fluoride, a competitive inhibitor of chloride activation. The results of these treatments will be examined using electron paramagnetic resonance (EPR) spectroscopy, a technique that is used to study molecules with unpaired electrons. A major objective of this project is to clarify the effects of calcium and chloride on the EPR signals from the oxygen evolving complex. To complement these studies, a second major objective is to use enzyme kinetics to characterize the interdependence of calcium and chloride in the activation of oxygen evolution.Broader ImpactThis project will be carried out in a department that has a major focus on involving undergraduate and Master's level graduate students in research. The university is a mid-sized public institution with a student population that is ethnically diverse and a traditional emphasis on undergraduate education. The university has recently experienced a growth in graduate programs, including a new PhD program in the department housing the project. The Principle Investigator has previously trained numerous beginning researchers, who have gone on to further graduate study or careers in industry, and the current project is designed to continue this type of training. The project involves a range of biochemical and biophysical techniques appropriate for various levels of experience. Beginning researchers can work fairly independently on experiments involving enzyme kinetics analyses and participate in the preparation and work-up of samples intended for EPR spectroscopy. Those more interested in the physical aspects of the research can participate directly in spectroscopic measurements and analysis. Overall, students gain exposure to both commonly used biochemical methods and more specialized spectroscopic techniques. Because of the great importance that photosynthetic processes have for supporting life in general, the presence on campus of a project that is focused on light energy conversion and oxygen production will help to raise awareness about energy utilization issues.