⁵¹V NMR Chemical Shifts Calculated from QM/MM Models of Vanadium Chloroperoxidase

Abstract

(51)V NMR chemical shifts calculated from QM/MM-optimized (QM=quantum mechanical; MM=molecular mechanical) models of vanadium-dependent chloroperoxidase (VCPO) are presented. An extensive number of protonation states for the vanadium cofactor (active site of the protein) and a number of probable positional isomers for each of the protonation states are considered. The size of the QM region is increased incrementally to observe the convergence behavior of the (51)V NMR chemical shifts. A total of 40 models are assessed by comparison to experimental solid-state (51)V NMR results recently reported in the literature. Isotropic chemical shifts are found to be a poor indicator of the protonation state; however, anisotropic chemical shifts and the nuclear quadrupole tensors appear to be sensitive to changes in the proton environment of the vanadium nuclei. This detailed investigation of the (51)V NMR chemical shifts computed from QM/MM models provides further evidence that the ground state is either a triply protonated (one axial water and one equatorial hydroxyl group) or a doubly protonated vanadate moiety in VCPO. Particular attention is given to the electrostatic and geometric effects of the protein environment. This is the first study to compute anisotropic NMR chemical shifts from QM/MM models of an active metalloprotein for direct comparison with solid-state MAS NMR data. This theoretical approach enhances the potential use of experimental solid-state NMR spectroscopy for the structural determination of metalloproteins.

51V NMR Chemical Shifts Calculated from QM/MM Models of Peroxo Forms of Vanadium Haloperoxidases

“…1.6-1.7 Å for vanadium oxo bonds (V=O), 1.8-1.9 Å for vanadium hydroxo (V-OH) and peroxo bonds (V-(O 2 )), and 2.0-2.1 Å for the V-N bond to HIS496.. As the standard deviations across the six snapshots for the bond lengths of the peroxovanadate cofactor are rather small, all the models are generally very similar within the six selected snapshots .The geometric convergence for the individual models is comparable to our prior QM/MM optimizations of the native VHPO forms, 22 indicating that the active site is rather rigid, and that the use of a small number of snapshots seems valid. Furthermore, a good degree of similarity is observed between the bond lengths of the peroxovanadate, across many of the different protonated models of p-VCPO and p-VBPO, see Tables 2 and S1b, respectively.…”

“…For instance, the three VCPO models that have reproduced the solid-state 51 V NMR-tensor elements reasonably well, VCPO-D14, -D34, and -T144, showed isotropic ( 51 V) values of -580, -607, and -602 ppm, respectively (largest QM region). 22 Except for the diprotonated p-VCPO-D13, all other p-VCPO models show clear upfield shifts from these numbers. For the p-VCPO-Z and p-VCPO-S1 variants, which emerged as particularly promising from the structural parameters discussed above, this computed upfield shift is between ca.…”

“…22,23 The SIMPSON 47 software package was employed to simulate the spectra, using the parameters extracted from the above CSA and EFG tensors. TD-DFT computations were performed at the same B3LYP/AE1+ level, for a number of different models, at the QM/MM optimized geometries.…”

“…22 Salient distances of all final p-VCPO models are summarized in Table 2, those of the corresponding p-VBPO homologues are deposited in Table S1b of the supporting information (SI).…”

Isomerization mechanism of the HcRed fluorescent protein chromophore

“…In the QM/MM approach, the ''active center'' and the environment are computed using an appropriate quantum mechanical method and a classical force field, respectively. This hybrid method enables a robust QM-based approach to the simulation of complex systems, 32,33 for example catalytic reactions of cytochrome P450 enzymes, 34,35 NMR chemical shifts of vanadium-dependent chloroperoxidase, 36 solvent effects on the electronic adsorption spectrum of guanine, 37 and photoactive yellow protein. 38 Our groups have previously studied the structural and energetic properties as well as the absorption spectra of some RFPs (DsRed.M1 and HcRed).…”