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• W2088682893 abstract "Escherichia coli ribonucleotide reductase is an α2β2 complex that catalyzes the conversion of nucleotides to deoxynucleotides using a diferric tyrosyl radical (Y122•) cofactor in β2 to initiate catalysis in α2. Each turnover requires reversible long-range proton-coupled electron transfer (PCET) over 35 Å between the two subunits by a specific pathway (Y122• ⇆ [W48?] ⇆ Y356 within β to Y731 ⇆ Y730 ⇆ C439 within α). Previously, we reported that a β2 mutant with 3-nitrotyrosyl radical (NO2Y•; 1.2 radicals/β2) in place of Y122• in the presence of α2, CDP, and ATP catalyzes formation of 0.6 equiv of dCDP and accumulates 0.6 equiv of a new Y• proposed to be located on Y356 in β2. We now report three independent methods that establish that Y356 is the predominant location (85–90%) of the radical, with the remaining 10–15% delocalized onto Y731 and Y730 in α2. Pulsed electron–electron double-resonance spectroscopy on samples prepared by rapid freeze quench (RFQ) methods identified three distances: 30 ± 0.4 Å (88% ± 3%) and 33 ± 0.4 and 38 ± 0.5 Å (12% ± 3%) indicative of NO2Y122•–Y356•, NO2Y122•–NO2Y122•, and NO2Y122•–Y731(730)•, respectively. Radical distribution in α2 was supported by RFQ electron paramagnetic resonance (EPR) studies using Y731(3,5-F2Y) or Y730(3,5-F2Y)-α2, which revealed F2Y•, studies using globally incorporated [β-2H2]Y-α2, and analysis using parameters obtained from 140 GHz EPR spectroscopy. The amount of Y• delocalized in α2 from these two studies varied from 6% to 15%. The studies together give the first insight into the relative redox potentials of the three transient Y• radicals in the PCET pathway and their conformations." @default.
• W2088682893 created "2016-06-24" @default.
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• W2088682893 date "2011-10-26" @default.
• W2088682893 modified "2023-10-03" @default.
• W2088682893 title "Equilibration of Tyrosyl Radicals (Y₃₅₆^•, Y₇₃₁^•, Y₇₃₀^•) in the Radical Propagation Pathway of the Escherichia coli Class Ia Ribonucleotide Reductase" @default.
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• W2088682893 doi "https://doi.org/10.1021/ja207455k" @default.
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