Pseudo‐Contact NMR Shifts over the Paramagnetic Metalloprotein CoMMP‐12 from First Principles

Abstract

Long-range pseudo-contact NMR shifts (PCSs) provide important restraints for the structure refinement of proteins when a paramagnetic metal center is present, either naturally or introduced artificially. Here we show that ab initio quantum-chemical methods and a modern version of the Kurland-McGarvey approach for paramagnetic NMR (pNMR) shifts in the presence of zero-field splitting (ZFS) together provide accurate predictions of all PCSs in a metalloprotein (high-spin cobalt-substituted MMP-12 as a test case). Computations of 314 13 C PCSs via g-and ZFS-tensors based on multi-reference methods provide a reliable bridge between EPRparameter-and susceptibility-based pNMR formalisms. Due to the high sensitivity of PCSs to even small structural differences, local structures based either on X-ray diffraction or on various DFT optimizations could be evaluated critically by comparing computed and experimental PCSs. Many DFT functionals provide insufficiently accurate structures. We also found the available 1RMZ PDB X-ray structure to exhibit deficiencies related to binding of a hydroxamate inhibitor. This has led to a newly refined PDB structure for MMP-12 (5LAB) that provides a more accurate coordination arrangement and PCSs.The anisotropic magnetic susceptibility [1] of paramagnetic metal ions induces the so-called pseudo-contact shifts (PCSs) in NMR spectra, which can be observed for nuclei between 5 Å and 40 Å from the metal center. [2] PCSs provide precious structural information on the biomolecules on which they are measured, both in solution and in the solid state. [3] PCS-based structural restraints have also become important for protein NMR crystallography. [4] Their importance is further enhanced by recent developments in fast magic-angle spinning (MAS) combined with high-field instruments. [4d, 5] While PCSs can thus provide crucial information on the structure of a metalloprotein as a whole, NMR is typically blind to nuclei near the paramagnetic metal center due to fast paramagnetic relaxation. The computation of pNMR shifts by first-principles quantum-chemical (QC) methods, on the other hand, has recently progressed appreciably, in particular by inclusion of the non-contact terms in small to medium-sized molecules, with no fundamental limitations close to the metal center. [6] There has so far been no attempt to access the longrange PCSs in larger biological systems by first-principles calculations, as the molecular sizes needed for an explicit treatment of the hyperfine coupling (HFC) anisotropies appeared prohibitive.Here we show that introduction of the point-dipole approximation (PDA), appropriate for the long-range spin-dipolar HFCs, into modern quantum-chemical pNMR shift machinery can be used to compute long-range PCSs based on accurate multireference ab initio calculations of g-and zero-field splitting (ZFS) D-tensors. Using such a combined approach, we have computed the entire set of 314 previously measured 13 C long-range PCSs [4a] (and further shifts from nuclei closer to the meta...

Pseudocontact shifts and paramagnetic susceptibility in semiempirical and quantum chemistry theories

“…essentially outside the room temperature thermal bath. 31 Since the submission of this manuscript, two important papers 37,38 and a review 39 were published dealing with the analysis of the pseudocontact shifts against the g and ZFS tensors, confirming the need of clarifying which relationship should be used to obtain the pseudocontact shifts from the EPR tensors. One paper, by Grandinetti and coworkers, 38 shows that the 2 H paramagnetic shifts measured for first row transition metal (manganese, iron, cobalt, nickel, or copper) chloride dihydrates can be well interpreted using the SE Kurland-McGarvey equation, i.e.…”

NMR Spectroscopy and Metal Ions in Life Sciences

“…Equation (2) which relates PCS to the magnetic susceptibility anisotropy of the metal center has been used since the sixties and has gone unchallenged until recently. In a collaborative work with theoretical groups, we have tested the possibility to predict PCS starting from first principles . This amounts to perform very accurate quantomechanical calculations, given an accurate 3D structure of the protein of interest and especially of the region encompassing the metal ion and its ligands.…”

Ab initio paramagnetic NMR shifts via point-dipole approximation in a large magnetic-anisotropy Co(ii) complex

“…7,8 Among the compounds of 3d transition metals, some cobalt complexes feature the combination of large zero-field splitting (ZFS) and g-tensor parameters that are commonly expected to translate to a large magnetic anisotropy. 9 Computational methods for the prediction and analysis of the magnetic properties of such paramagnetic systems have progressed during recent years. The NMR chemical shifts induced by a strongly anisotropic magnetic center have already for a long time been calculated by a longdistance PDA of the dipolar hyperfine coupling (HFC) interactions between the unpaired electrons and NMR nuclei.…”