Spectroscopic studies of light-driven electron transfer

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Long distance electron transfer plays an important role in biological function in many enzymes. In these reactions, radical transport may occur and may involve transfer of an electron alone (ET) or a proton coupled, electron transfer reaction (PET). The mechanism by which the protein environment controls these reactions is just beginning to be elucidated. PET reactions in ribonucleotide reductase (RNR) and photosystem II (PSII) are the immediate focus of this application. PSII carries out the light-induced oxidation of water and reduction of plastoquinone. In PSII, a redox-active tyrosine, YZ, conducts electrons between the manganese-containing oxygen-evolving center (GEC) and the primary electron donor. A second redox-active tyrosine, YO, is oxidized by the primary electron donor, but is not required for oxygen-evolving activity. RNR catalyzes the reduction of ribonucleotides to deoxynucleotides. In E. coli RNR, a redox-active tyrosine, Y122, is proposed to be a radical initiator. In this proposal, spectroscopic methods will be employed, which will identify radical intermediates and elucidate catalytic mechanism in these two proteins. Studies will also be conducted of peptide maquettes, which will be used to test hypotheses generated from studies of the natural systems. This proposal has three specific aims. In A, we will use transient EPR and infrared spectroscopies to test the hypothesis that the mechanism of proton-coupled electron transfer distinguishes the redox-active tyrosines, YO and YZ, in PSII. Because the two PSII tyrosines have different redox and kinetic properties, new information will be acquired concerning the functional control of biological PET reactions. In B, we will employ a new method, stopped flow FT-IR spectroscopy, to probe the mechanism of proton-coupled electron transfer in RNR. We will test the hypothesis that redox changes at Y122 are coupled with structural changes in adjacent peptide bonds. In C, we will use designed beta hairpin maquettes to test the hypotheses that proton-coupled electron transfer can occur from tyrosine to a pi-pi stacked histidine. This specific aim will serve to elucidate the role of primary, secondary, and tertiary interactions in a structurally defined, tractable model system.