ArticleJournal of biomolecular NMR2001
Calmodulin tagging provides a general method of using lanthanide induced magnetic field orientation to observe residual dipolar couplings in proteins in solution.
Article in Journal of biomolecular NMR, 2001. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
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18 citing papers in PubMed, 54 citations in OpenAlex.
- Synthesis of lanthanide tag and experimental studies on paramagnetically induced residual dipolar couplings.BMC chemistry · 2022Article
- Localization of ligands within human carbonic anhydrase II usingChemical science · 2019Article
- Long-Range RNA Structural Information via a Paramagnetically Tagged Reporter Protein.Journal of the American Chemical Society · 2019Article
- An encodable lanthanide binding tag with reduced size and flexibility for measuring residual dipolar couplings and pseudocontact shifts in large proteins.Journal of biomolecular NMR · 2016Article
- Multiple paramagnetic effects through a tagged reporter protein.Angewandte Chemie (International ed. in English) · 2015Article
- Engineering encodable lanthanide-binding tags into loop regions of proteins.Journal of the American Chemical Society · 2011Article
- Integrated analysis of the conformation of a protein-linked spin label by crystallography, EPR and NMR spectroscopy.Journal of biomolecular NMR · 2011Article
- Paramagnetic labelling of proteins and oligonucleotides for NMR.Journal of biomolecular NMR · 2010Review
- Hyperfine-shifted (13)C and (15)N NMR signals from Clostridium pasteurianum rubredoxin: extensive assignments and quantum chemical verification.Journal of the American Chemical Society · 2009Article
- Phage-induced alignment of membrane proteins enables the measurement and structural analysis of residual dipolar couplings with dipolar waves and lambda-maps.Journal of the American Chemical Society · 2009Article
- High-resolution structure determination of the CylR2 homodimer using paramagnetic relaxation enhancement and structure-based prediction of molecular alignment.Journal of biomolecular NMR · 2008Article
- Bound-state residual dipolar couplings for rapidly exchanging ligands of His-tagged proteins.Journal of the American Chemical Society · 2007Article
- Multiple alignment of membrane proteins for measuring residual dipolar couplings using lanthanide ions bound to a small metal chelator.Journal of the American Chemical Society · 2007Article
- Novel techniques for weak alignment of proteins in solution using chemical tags coordinating lanthanide ions.Journal of biomolecular NMR · 2004Article
- Improving the accuracy of NMR structures of large proteins using pseudocontact shifts as long-range restraints.Journal of biomolecular NMR · 2004Article
- Using lanthanide ions to align troponin complexes in solution: order of lanthanide occupancy in cardiac troponin C.Protein science : a publication of the Protein Society · 2004Article
- Breaking symmetry in the structure determination of (large) symmetric protein dimers.Journal of biomolecular NMR · 2002Article
- Structure-independent cross-validation between residual dipolar couplings originating from internal and external orienting media.Journal of biomolecular NMR · 2002Article
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5 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
A general method is presented for magnetic field alignment of proteins in solution. By tagging a target protein with calmodulin saturated with paramagnetic lanthanide ions it is possible to measure substantial residual dipolar couplings (RDC) whilst minimising the effects of pseudocontact shifts on the target protein. A construct was made consisting of a calmodulin-binding peptide (M13 from sk-MLCK) attached to a target protein, dihydrofolate reductase in this case. The engineered protein binds tightly to calmodulin saturated with terbium, a paramagnetic lanthanide ion. By using only a short linker region between the M13 and the target protein, some of the magnetic field alignment induced in the CaM(Tb3+)4 is effectively transmitted to the target protein (DHFR). 1H-15N HSQC IPAP experiments on the tagged complex containing 15N-labelled DHFR-M13 protein and unlabelled CaM(Tb3+)4 allow one to measure RDC contributions in the aligned complex. RDC values in the range +4.0 to -7.4 Hz were measured at 600 MHz. Comparisons of 1H-15N HSQC spectra of 15N-DHFR-M13 alone and its complexes with CaM(Ca2+)4 and CaM(Tb3+)4 indicated that (i) the structure of the target protein is not affected by the complex formation and (ii) the spectra of the target protein are not seriously perturbed by pseudocontact shifts. The use of a relatively large tagging group (CaM) allows us to use a lanthanide ion with a very high magnetic susceptibility anisotropy (such as Tb3+) to give large alignments while maintaining relatively long distances from the target protein nuclei (and hence giving only small pseudocontact shift contributions).
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