J. Am. Chem. Soc., 128 (15), 5190 -5208, 2006. 10.1021/ja060480f S0002-7863(06)00480-X
51V Solid-State Magic Angle Spinning NMR Spectroscopy of Vanadium Chloroperoxidase
Neela Pooransingh-Margolis, Rokus Renirie, Zulfiqar Hasan, Ron Wever, Alexander J. Vega, and Tatyana Polenova*
Contribution from the Brown Laboratories, Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716, Van 't Hoff Institute for Molecular Sciences, Faculty of Science, University of Amsterdam, NieuweAchtergracht 129, 1018 WS Amsterdam, The Netherlands
Received January 20, 2006
We report 51V solid-state NMR spectroscopy of the 67.5-kDa vanadium chloroperoxidase, at 14.1 T. We demonstrate that, despite the low concentration of vanadium sites in the protein (one per molecule, 1 mol of vanadium spins in the entire sample), the spinning sideband manifold spanning the central and the satellite transitions is readily detectable. The quadrupolar and chemical shift anisotropy tensors have been determined by numerical simulations of the spinning sideband envelopes and the line shapes of the individual spinning sidebands corresponding to the central transition. The observed quadrupolar coupling constant CQ of 10.5 ± 1.5 MHz and chemical shift anisotropy of -520 ± 13 ppm are sensitive reporters of the geometric and electronic structure of the vanadium center. Density functional theory calculations of the NMR spectroscopic observables for an extensive series of active site models indicate that the vanadate cofactor is most likely anionic with one axial hydroxo- group and an equatorial plane consisting of one hydroxo- and two oxo- groups. The work reported in this manuscript is the first example of 51V solid-state NMR spectroscopy applied to probe the vanadium center in a protein directly. This approach yields the detailed coordination environment of the metal unavailable from other experimental measurements and is expected to be generally applicable for studies of diamagnetic vanadium sites in metalloproteins.