The elucidation of the biosynthetic pathway of the molybdenum cofactor has enabled direct incorporation of 33S-labeled sulfide, using controlled in vitro synthesis with purified proteins, to produce a catalytically active variant of sulfite oxidase (SO). Pulsed EPR spectroscopy of its Mo(V) state at low pH has provided the first direct measurements of the interaction of an unpaired electron on a metal center with the coordinating S atoms of a dithiolene ligand. Numerical simulations show that the 33S ESEEM spectra are dominated by the 'inter-doublet' transition arising from strong nqi, as has been observed previously for 33S-labeled sulfite bound to SO. The estimated experimental values for the 33S spectroscopic parameters are aiso = 3 MHz and e2Qq/h = 25 MHz. The DFT calculations show spin polarization effects and strong intermixing between orbitals with Mo dxy character and the out-of-plane pz orbitals of the MPT S atoms, providing a possible mechanism for significant 33S hfianisotropy. Additionally, this intermixing may be an important factor in the proposed role of the dithiolene unit of the cofactor in effective coupling into protein-mediated superexchange pathways for efficient electron transfer during catalysis.

Insights into metal dithiolene electronic structure from pulsed EPR spectroscopy and DFT calculations on <sup>33</sup>S-labeled molybdenum cofactor in catalytically active bioengineered sulfite oxidase

248th National Meeting (2014)

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Insights into metal dithiolene electronic structure from pulsed EPR spectroscopy and DFT calculations on 33S-labeled molybdenum cofactor in catalytically active bioengineered sulfite oxidase

Insights into metal dithiolene electronic structure from pulsed EPR spectroscopy and DFT calculations on ³³S-labeled molybdenum cofactor in catalytically active bioengineered sulfite oxidase