Intermolecular dynamics studied by paramagnetic tagging

Xu, Xiaozheng; Keizers, Peter H. J.; Reinle, Wolfgang; Hannemann, Frank; Bernhardt, Rita; Ubbink, Marcellus

doi:10.1007/s10858-009-9308-0

Cited by 56 publications

(42 citation statements)

References 42 publications

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“…The structurally highly similar tag Ln-CLaNP-5 shows a single set of resonances when attached to the same double Cys mutant of Cyt c ( Figure S5). 33 Also for CLaNP-7 attached to Paz E51C/E54C no pH dependence of the PCS was observed, so the observed effect is specific for CLaNP-7 attached to Cyt c N56C/L58C. Protonation of the phenol moiety of CLaNP-7 as an explanation for the pH dependence was ruled out by the electronic absorption spectrum.…”

Section: ■ Resultsmentioning

confidence: 92%

A pH-Sensitive, Colorful, Lanthanide-Chelating Paramagnetic NMR Probe

et al. 2012

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Paramagnetic lanthanides ions are broadly used in NMR spectroscopy. The effects of unpaired electrons on NMR spectral parameters provide a powerful tool for the characterization of macromolecular structures and dynamics. Here, a new lanthanide-chelating NMR probe, Caged Lanthanide NMR Probe-7 (CLaNP-7), is presented. It can be attached to protein surfaces via two disulfide bridges, yielding a probe that is rigid relative to the protein backbone. CLaNP-7 extends the application range of available probes. It has a yellow color, which is helpful for sample preparation. Its effects are comparable to those of CLaNP-5, but its charge is two units lower (+1) than that of CLaNP-5 (+3), reducing the change in surface potential after probe attachment. It also has a different magnetic susceptibility tensor, so by using both tags, two sets of structural restraints can be obtained per engineered cysteine pair. Moreover, it was found that the orientation of the magnetic susceptibility tensor is pH dependent (pK(a) ≈ 7) when a histidine residue is located in the neighborhood of the probe attachment site. The results show that the His imidazole group interacts with the CLaNP-7 tag. It is proposed that the histidine residue forms a hydrogen bond to a water/hydroxyl molecule that occupies the ninth coordination position on the lanthanide, thus breaking the two-fold symmetry of the CLaNP tag in a pH-dependent way.

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Section: ■ Resultsmentioning

confidence: 92%

A pH-Sensitive, Colorful, Lanthanide-Chelating Paramagnetic NMR Probe

et al. 2012

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“…Therefore, spin labeling or the use of paramagnetic tags is required to exploit the powerful paramagnetic restraints described above. Various spin labels with stabilized nitroxyl radicals and paramagnetic lanthanide binding tags are available and can be attached to single cysteine residues (after removing additional cysteines by site-directed mutagenesis) in a domain or subunit of a protein complex (Battiste and Wagner, 2000;Gaponenko et al, 2000;Clore and Iwahara, 2009;Xu et al, 2009;Simon et al, 2010;Su and Otting, 2010a).…”

Section: Use Of Paramagnetism To Provide Long-range Distance (Pre) Anmentioning

confidence: 99%

NMR and small-angle scattering-based structural analysis of protein complexes in solution

Madl

Gabel

Sattler

2011

Journal of Structural Biology

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“…One technique that has gained popularity in recent years is the site-specific incorporation of paramagnetic spin labels or lanthanide binding tags. [3] These labels are covalently attached to single cysteine residues and provide a rich source of distance (paramagnetic relaxation enhancements) and orientation information (pseudocontact shifts, PCS; residual dipolar couplings, RDCs). [3a,b,d] A potential drawback of this approach is the requirement of a single accessible cysteine residue for cross-linking with the paramagnetic tag.…”

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confidence: 99%