Paramagpy: software for fitting magnetic susceptibility tensors using paramagnetic effects measured in NMR spectra

Orton, Henry; Huber, Thomas; Otting, Gottfried

doi:10.5194/mr-1-1-2020

Cited by 41 publications

(46 citation statements)

References 41 publications

(103 reference statements)

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“…Figure 1a shows the PCSs observed with Tb 3+ and Tm 3+ ions. Using the NMR ensemble structure of ubiquitin (PDB ID: 2KOX; Fenwick et al, 2011) and the measured PCSs, the metal position was determined by fitting the χ tensor using the program Paramagpy (Table 1; Orton et al, 2020). The correlation between back-calculated and experimental PCSs was excellent ( Fig.…”

Section: Single Phosphoserine Residues For Lanthanide Bindingmentioning

confidence: 95%

“…PCSs were used to determine the position and orientation of the χ tensor of the paramagnetic ions relative to the protein structure. Fitting of χ tensors was performed using the program Paramagpy (Orton et al, 2020).…”

Section: Pcs Measurements and ∆χ -Tensor Fittingmentioning

confidence: 99%

“…Paramagnetic labels offer an attractive tool for the study of protein structure and function, as the magnetic moments of unpaired electrons generate long-range paramagnetic effects in nuclear magnetic resonance (NMR) spectra. Among the paramagnetic effects that can be observed in NMR spectra, pseudocontact shifts (PCSs) generated by paramagnetic metal ions stand out for their high information content and ease of observation (Otting, 2008;Parigi and Luchinat, 2018). Specifically, the PCSs provide information about the location of nuclear spins relative to the magnetic susceptibility anisotropy tensor ( χ tensor) associated with a paramagnetic metal ion, and this information can readily be obtained for nuclear spins as far as 40 Å from the paramagnetic centre (Bertini et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

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Phosphoserine for the generation of lanthanide-binding sites on proteins for paramagnetic nuclear magnetic resonance spectroscopy

et al. 2021

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Abstract. Pseudocontact shifts (PCSs) generated by paramagnetic lanthanide ions provide valuable long-range structural information in nuclear magnetic resonance (NMR) spectroscopic analyses of biological macromolecules such as proteins, but labelling proteins site-specifically with a single lanthanide ion remains an ongoing challenge, especially for proteins that are not suitable for ligation with cysteine-reactive lanthanide complexes. We show that a specific lanthanide-binding site can be installed on proteins by incorporation of phosphoserine in conjunction with other negatively charged residues, such as aspartate, glutamate or a second phosphoserine residue. The close proximity of the binding sites to the protein backbone leads to good immobilization of the lanthanide ion, as evidenced by the excellent quality of fits between experimental PCSs and PCSs calculated with a single magnetic susceptibility anisotropy (Δχ) tensor. An improved two-plasmid system was designed to enhance the yields of proteins with genetically encoded phosphoserine, and good lanthanide ion affinities were obtained when the side chains of the phosphoserine and aspartate residues are not engaged in salt bridges, although the presence of too many negatively charged residues in close proximity can also lead to unfolding of the protein. In view of the quality of the Δχ tensors that can be obtained from lanthanide-binding sites generated by site-specific incorporation of phosphoserine, this method presents an attractive tool for generating PCSs in stable proteins, particularly as it is independent of cysteine residues.

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Section: Single Phosphoserine Residues For Lanthanide Bindingmentioning

confidence: 95%

Section: Pcs Measurements and ∆χ -Tensor Fittingmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Phosphoserine for the generation of lanthanide-binding sites on proteins for paramagnetic nuclear magnetic resonance spectroscopy

et al. 2021

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“…62 The PCS can then be translated into structural restraints by fitting of the experimental data to a known or calculated structure using software packages like Numbat 97 or Paramagpy. 98 Programs capable of PCS analysis minimize the difference between observed and back-calculated PCS in an iterative fashion until the convergence is achieved. The resulting axial and rhombic parameters of the observed anisotropy tensors should be reported in the unique tensor representation (UTR) using the zyz' convention, as only this representation allows for direct comparison with tensors from other LCTs.…”

Section: Paramagnetic Nuclear Magnetic Resonance Spectroscopymentioning

confidence: 99%

“…As discussed, software packages, e.g. Numbat 97 or Paramagpy 98 are used to derive the anisotropy parameters from the experimental data by minimizing the difference between the experimental and back-calculated PCS for a given structure (Fig. 5).…”

Section: Pcs Polar Coordinatesmentioning

confidence: 99%

Application of Paramagnetic Lanthanoid Chelating Tags in NMR Spectroscopy and Their Use for the Localization of Ligands Within Biomacromolecules

Joss

Vogel

Zimmermann

et al. 2021

Comprehensive Coordination Chemistry III

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DOTA-Based Lanthanoid Chelating Tags 8 3 Applications of Lanthanoid Chelating Tags in the Localization of Ligands Within Biomacromolecules 12 3.1 Introduction 12 3.2 Weakly Binding Ligands 12 3.3 Ligands in the Intermediate Exchange Regime 13 3.4 Strongly Binding Ligands 14 4 Localization of Ligands Within Human Carbonic Anhydrase II Using 19 F Pseudocontact Shift Analysis 15 4.1 Human Carbonic Anhydrase and Its Use in the Development of Artificial Metalloenzymes 15 4.2 Localization of Ligands within Human Carbonic Anhydrase II-Introduction and Concept 16 4.3 PCS Analysis and Determination of the Anisotropy Parameters from Multiple Positions Within the Protein 18 4.4 Measurement of 19 F PCS and Localization of the Ligand Using a Monte-Carlo Approach 19 4.5 Determination of the Ligand Position by Analysis of the Isosurfaces of the Anisotropy Parameters 24 4.6 1 H-19 F-HOESY Experiments for Determination of 1 H PCS of the Ligand and Protein-Ligand Interactions 24 5 Conclusion and Outlook 26 Acknowledgment 26 References 26

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A Chiral Lanthanide Tag for Stable and Rigid Attachment to Single Cysteine Residues in Proteins for NMR, EPR and Time‐Resolved Luminescence Studies

Herath

Breen

Hewitt

et al. 2021

Chemistry A European J

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A lanthanide-binding tag site-specifically attached to a protein presents a tool to probe the protein by multiple spectroscopic techniques, including nuclear magnetic resonance, electron paramagnetic resonance and time-resolved luminescence spectroscopy. Here a new stable chiral Ln III tag, referred to as C12, is presented for spontaneous and quantitative reaction with a cysteine residue to generate a stable thioether bond. The synthetic protocol of the tag is relatively straightforward, and the tag is stable for storage and shipping. It displays greatly enhanced reactivity towards selenocysteine, opening a route towards selective tagging of selenocysteine in proteins containing cysteine residues.Loaded with Tb III or Tm III ions, the C12 tag readily generates pseudocontact shifts (PCS) in protein NMR spectra. It produces a relatively rigid tether between lanthanide and protein, which is beneficial for interpretation of the PCSs by single magnetic susceptibility anisotropy tensors, and it is suitable for measuring distance distributions in double electron-electron resonance experiments. Upon reaction with cysteine or other thiol compounds, the Tb III complex exhibits a 100-fold enhancement in luminescence quantum yield, affording a highly sensitive turn-on luminescence probe for time-resolved FRET assays and enzyme reaction monitoring.

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Paramagpy: software for fitting magnetic susceptibility tensors using paramagnetic effects measured in NMR spectra

Cited by 41 publications

References 41 publications

Phosphoserine for the generation of lanthanide-binding sites on proteins for paramagnetic nuclear magnetic resonance spectroscopy

Phosphoserine for the generation of lanthanide-binding sites on proteins for paramagnetic nuclear magnetic resonance spectroscopy

Application of Paramagnetic Lanthanoid Chelating Tags in NMR Spectroscopy and Their Use for the Localization of Ligands Within Biomacromolecules

A Chiral Lanthanide Tag for Stable and Rigid Attachment to Single Cysteine Residues in Proteins for NMR, EPR and Time‐Resolved Luminescence Studies

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