Model-free extraction of spin label position distributions from pseudocontact shift data

Suturina, Elizaveta A.; Häußinger, Daniel; Zimmermann, Kaspar; Garbuio, Luca; Yulikov, Maxim; Jeschke, Gunnar; Kuprov, Ilya

doi:10.1039/c6sc03736d

Cited by 26 publications

(29 citation statements)

References 48 publications

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“…In order to investigate the mobility of the newly developed lanthanide‐chelating tag on the protein surface, the metal centre distribution was modelled using a method developed by Suturina et al. (Figure ) . In PCS NMR spectroscopy, the point paramagnetic centre approximation is usually applied.…”

Section: Resultsmentioning

confidence: 99%

“…1 H‐ 15 N HSQC in‐phase/anti‐phase (IPAP) spectra were measured from solutions in 10 m m phosphate buffer at pH 6.0 and 298 K on a 900 MHz Bruker Avance III NMR spectrometer equipped with a TXI probe (ubiquitin S57 C) or in 10 m m phosphate buffer at pH 6.8 and 298 K on a 600 MHz Bruker Avance III NMR spectrometer equipped with a QCI‐F probe (hCA II S166 C). Assignments of the obtained NMR spectra were made using CcpNmr Analysis . The tensor properties were then obtained by fitting to the residues in secondary structure elements of ubiquitin (PDB 1UBI 37 ) or the leucine residues of hCA II (PDB 3KS3) using Numbat .…”

Section: Methodsmentioning

confidence: 99%

“…In order to investigate the mobility of the newly developed lanthanide-chelatingt ag on the protein surface, the metal centre distribution was modelled using am ethod developed by Suturina et al ( Figure1 1). [33][34][35] In PCS NMR spectroscopy, the point paramagnetic centre approximation is usually applied. Since the protein is measured in solution at room temperaturea nd LCTsm aintain their flexibility,t he metal position can be more realistically described in terms of its probability density.T he approach developed by Suturinae tal.…”

Section: Properties Of the Isopropyl-substituted Lanthanidechelating mentioning

confidence: 99%

“…Assignments of the obtained NMR spectra were made using CcpNmr Analysis. [33,39,40] The tensor properties were then obtained by fitting to the residues in secondary structure elements of ubiquitin (PDB 1UBI 37 )o rt he leucine residues of hCA II (PDB 3KS3 [41] )u sing Numbat. [42] Q-factors were calculated according to the standard equation described by Nitsche et al [1] DFT calculations DFT calculations were performed with the ORCA program package [43] at the sciCORE facility of the University of Basel.…”

Section: Expression Of Ubiquitin S57 Ca Nd Tagging Reactionmentioning

confidence: 99%

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A Sterically Overcrowded, Isopropyl‐Substituted, Lanthanide‐Chelating Tag for Protein Pseudocontact Shift NMR Spectroscopy: Synthesis of its Macrocyclic Scaffold and Benchmarking on Ubiquitin S57 C and hCA II S166 C

Joss

Bertrams

Häußinger

2019

Chemistry A European J

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A sterically overcrowded lanthanide‐chelating tag has been synthesized in order to investigate the influence on the obtained pseudocontact shifts and the anisotropic part of the magnetic susceptibility tensor compared to those of its predecessor DOTA‐M8‐(4R,4S)‐SSPy. For the first time, a concise synthetic route is presented for isopropyl‐substituted cyclen, the macrocyclic scaffold of the lanthanide‐chelating tag, delivering the macrocycle in an overall yield of 6 % over 11 steps. The geometry of the lutetium complex has been assigned by ROESY experiments, adopting exclusively a Λ(δδδδ) conformation, and DFT calculations have confirmed a stabilization of 32.6 kJ mol−1 compared to the Δ(δδδδ) conformer. The highly rigidified lanthanide‐chelating tag induces strong pseudocontact shifts of up to 6.5 ppm on ubiquitin S57 C, shows significantly improved tensor properties compared to those of its predecessor, and constitutes a highly promising starting point for the further development of lanthanide‐chelating tags.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Properties Of the Isopropyl-substituted Lanthanidechelating mentioning

confidence: 99%

Section: Expression Of Ubiquitin S57 Ca Nd Tagging Reactionmentioning

confidence: 99%

See 2 more Smart Citations

A Sterically Overcrowded, Isopropyl‐Substituted, Lanthanide‐Chelating Tag for Protein Pseudocontact Shift NMR Spectroscopy: Synthesis of its Macrocyclic Scaffold and Benchmarking on Ubiquitin S57 C and hCA II S166 C

Joss

Bertrams

Häußinger

2019

Chemistry A European J

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“…In order to compare the experimental distances with the proteins’ structures, softwares are available to attach in silico a spin label to an existing PDB structure and simulate the most probable inter‐spin distances using rotamer libraries. Here we provide new rotamer libraries for gem ‐diethyl labels compatible with the MMM software, generated by an extended version of the approach previously used for other nitroxide and for Gd(III) labels …”

Section: New Rotamer Libraries and Interspin Distance Simulationsmentioning

confidence: 99%

gem‐Diethyl Pyrroline Nitroxide Spin Labels: Synthesis, EPR Characterization, Rotamer Libraries and Biocompatibility

et al. 2019

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The availability of bioresistant spin labels is crucial for the optimization of site‐directed spin labeling protocols for EPR structural studies of biomolecules in a cellular context. As labeling can affect proteins’ fold and/or function, having the possibility to choose between different spin labels will increase the probability to produce spin‐labeled functional proteins. Here, we report the synthesis and characterization of iodoacetamide‐ and maleimide‐functionalized spin labels based on the gem ‐diethyl pyrroline structure. The two nitroxide labels are compared to conventional gem ‐dimethyl analogs by site‐directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy, using two water soluble proteins: T4 lysozyme and Bid. To foster their use for structural studies, we also present rotamer libraries for these labels, compatible with the MMM software. Finally, we investigate the “true” biocompatibility of the gem ‐diethyl probes comparing the resistance towards chemical reduction of the NO group in ascorbate solutions and E. coli cytosol at different spin concentrations.

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Unravelling the Complexities of Pseudocontact Shift Analysis in Lanthanide Coordination Complexes of Differing Symmetry

et al. 2019

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In two closely related series of eight-coordinate lanthanide complexes,aswitch in the sign of the dominant ligand field parameter and striking variations in the sign, amplitude and orientation of the main component of the magnetic susceptibility tensor as the Ln 3+ ion is permuted conspire to mask modest changes in NMR paramagnetic shifts, but are evident in Yb EPR and Eu emission spectra.Lanthanide induced shift and paramagnetic relaxation enhancement are subjects of continuous research driven by new applications of lanthanide complexes as tags for biomolecular structure analysis, [1] and PA RASHIFT probes for magnetic resonance imaging (MRI). [2] While Gd 3+ tags are employed in double electron-electron resonance (DEER) distance measurements in biomolecules, [3] and Gd 3+ MRI contrast agents are routinely used, [4] non-Gd tags with pronounced magnetic anisotropy continue to challenge existing theoretical models of paramagnetic shift [5] and relaxation. [6] Many of the conclusions made in the early literature examining NMR behaviour in paramagnetic lanthanide systems,f or example,m ay have low significance because of the limitations of Bleaneyst heory of magnetic anisotropy. [7] Often, unreasonable estimations of ac ontact contribution were invoked in order to allow experimental data to be fitted, and sometimes "distance only" dependent relaxation rate data were used to aid assignment, notwithstanding the fact that the electron-nuclear dipolar interaction has been shown to have as trong directional dependence,b oth in the dipolar and the Curie contributions to paramagnetic relaxation. [6] Thet heory of Bleaney assumed that ligand field (zerofield) splitting of the ground J-multiplet is less than kT and that the ligand-field parameters are invariant as the lanthanide ion is permuted in an isostructural series.N either Supportinginformation and the ORCID identification number(s) for the author(s) of this article can be found under https://doi.

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Model-free extraction of spin label position distributions from pseudocontact shift data

Abstract: Not a point, but a cloud: advanced PCS data analysis using 3D probability density reconstruction provides more information.

Cited by 26 publications

References 48 publications

A Sterically Overcrowded, Isopropyl‐Substituted, Lanthanide‐Chelating Tag for Protein Pseudocontact Shift NMR Spectroscopy: Synthesis of its Macrocyclic Scaffold and Benchmarking on Ubiquitin S57 C and hCA II S166 C

A Sterically Overcrowded, Isopropyl‐Substituted, Lanthanide‐Chelating Tag for Protein Pseudocontact Shift NMR Spectroscopy: Synthesis of its Macrocyclic Scaffold and Benchmarking on Ubiquitin S57 C and hCA II S166 C

gem‐Diethyl Pyrroline Nitroxide Spin Labels: Synthesis, EPR Characterization, Rotamer Libraries and Biocompatibility

Unravelling the Complexities of Pseudocontact Shift Analysis in Lanthanide Coordination Complexes of Differing Symmetry

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