Geometric and electronic structure of the Mn(IV)Fe(III) cofactor in class Ic ribonucleotide reductase: Correlation to the class ia binuclear non-heme iron enzyme

The class Ic ribonucleotide reductase (RNR) from Chlamydia trachomatis (Ct) utilizes a Mn/Fe heterobinuclear cofactor, rather than the Fe/Fe cofactor found in the β (R2) subunit of the class Ia enzymes, to react with O ₂. This reaction produces a stable Mn^IVFe^III cofactor that initiates a radical, which transfers to the adjacent α (R1) subunit and reacts with the substrate. We have studied the Mn ^IVFe^III cofactor using nuclear resonance vibrational spectroscopy (NRVS) and absorption (Abs)/circular dichroism (CD)/magnetic CD (MCD)/variable temperature, variable field (VTVH) MCD spectroscopies to obtain detailed insight into its geometric/electronic structure and to correlate structure with reactivity; NRVS focuses on the Fe^III, whereas MCD reflects the spin-allowed transitions mostly on the Mn^IV. We have evaluated 18 systematically varied structures. Comparison of the simulated NRVS spectra to the experimental data shows that the cofactor has one carboxylate bridge, with Mn^IV at the site proximal to Phe₁₂₇. Abs/CD/MCD/VTVH MCD data exhibit 12 transitions that are assigned as d-d and oxo and OH^- to metal charge-transfer (CT) transitions. Assignments are based on MCD/Abs intensity ratios, transition energies, polarizations, and derivative-shaped pseudo-A term CT transitions. Correlating these results with TD-DFT calculations defines the Mn^IVFe^III cofactor as having a μ-oxo, μ-hydroxo core and a terminal hydroxo ligand on the Mn^IV. From DFT calculations, the Mn^IV at site 1 is necessary to tune the redox potential to a value similar to that of the tyrosine radical in class Ia RNR, and the OH^- terminal ligand on this Mn ^IV provides a high proton affinity that could gate radical translocation to the α (R1) subunit.