Modulation of Y₃₅₆ photooxidation in E. coli class Ia ribonucleotide reductase by Y₇₃₁ across the α₂: β₂ interface

Substrate turnover in class Ia ribonucleotide reductase (RNR) requires reversible radical transport across two subunits over 35 Å, which occurs by a multistep proton-coupled electron-transfer mechanism. Using a photooxidant-labeled β₂ subunit of Escherichia coli class Ia RNR, we demonstrate photoinitiated oxidation of a tyrosine in an α₂:β₂ complex, which results in substrate turnover. Using site-directed mutations of the redox-active tyrosines at the subunit interface, Y₃₅₆F(β) and Y₇₃₁F(α), this oxidation is identified to be localized on Y₃₅₆. The rate of Y ₃₅₆ oxidation depends on the presence of Y₇₃₁ across the interface. This observation supports the proposal that unidirectional PCET across the Y₃₅₆(β)-Y₇₃₁(α)-Y ₇₃₀(α) triad is crucial to radical transport in RNR.