In the broadest sense, our objective is to understand biological processes and phenomena in terms of physical mechanisms, i.e. on a molecular level. The main problem addressed in this proposal deals with the conversion of electromagnetic energy (light) into chemical energy, i.e. photosynthesis. This process is mediated by an integral membrane protein - pigment complex called the reaction center (RC). Our work focuses on the structural aspects of the RC from the photosynthetic bacterium Rb. sphaeroides and the structural implications on the function of the RC. The mechanisms of electron and proton transfers that take place in the RC are of general importance in all living systems. For instance, the processes that take place in mitochondria (in mammalian cells) can be viewed as the reverse process of photosynthesis utilizing similar mechanisms. Thus, the understanding of electron and proton transfer mechanisms should lead to a deeper understanding of the molecular basis of some metabolic deficiencies. The fields and methodologies used in this proposal are interdisciplinary, ranging from physics to chemistry and molecular biology. Consequently, different techniques and tools are used in our work, e.g. computational methods, X-ray crystallography, optical spectroscopy, magnetic resonance (EPR, ENDOR), protein chemistry and recombinant DNA techniques. The specific problems addressed are: The three-dimensional structure of native and mutant RCs from the photosynthetic bacterium Rb. sphaeroides and the structure of the transient RC-cyt c2 complex. These studies are being pursued by X-ray crystallography, complemented by magnetic resonance techniques. The mechanisms of two electron transfer reactions: k(1)AB(Q-AQB->QAQ-B) and k(2)AB (Q-AQ-B+H+->Q(QBH)-), as well as the protonation of the secondary quinone, QB, are being studied by kinetic optical spectroscopy and site-directed mutagenesis.
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