Three-Dimensional Protein Fold Determination from Backbone Amide Pseudocontact Shifts Generated by Lanthanide Tags at Multiple Sites
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AbstractSite-specific attachment of paramagnetic lanthanide ions to a protein generates pseudocontact shifts (PCS) in the nuclear magnetic resonance (NMR) spectra of the protein that are easily measured as changes in chemical shifts. By labeling the protein with lanthanide tags at four different sites, PCSs are observed for most amide protons and accurate information is obtained about their coordinates in three-dimensional space. The approach is demonstrated with the chaperone ERp29, for which large differences have been reported between X-ray and NMR structures of the C-terminal domain, ERp29-C. The results unambiguously show that the structure of rat ERp29-C in solution is similar to the crystal structure of human ERp29-C. PCSs of backbone amides were the only structural restraints required. Because these can be measured for more dilute protein solutions than other NMR restraints, the approach greatly widens the range of proteins amenable to structural studies in solution. © 2013 Elsevier Ltd. All rights reserved.
CitationYagi H, Pilla KB, Maleckis A, Graham B, Huber T, et al. (2013) Three-Dimensional Protein Fold Determination from Backbone Amide Pseudocontact Shifts Generated by Lanthanide Tags at Multiple Sites. Structure 21: 883–890. Available: http://dx.doi.org/10.1016/j.str.2013.04.001.
SponsorsWe thank Dr. Souren Mkrtchian for the plasmid encoding ERp29 and the supercomputing facility at the King Abdulla University of Science and Technology (KAUST, Saudi Arabia) for providing access to the Blue Gene/P (Shaheen) supercomputer. Financial support by the Australian Research Council, including a Future Fellowship to T.H., is gratefully acknowledged.
CollectionsPublications Acknowledging KAUST Support
- Three-Dimensional Protein Structure Determination Using Pseudocontact Shifts of Backbone Amide Protons Generated by Double-Histidine Co<sup>2+</sup>-Binding Motifs at Multiple Sites.
- Authors: Bahramzadeh A, Huber T, Otting G
- Issue date: 2019 Jul 30
- Generation of pseudocontact shifts in proteins with lanthanides using small "clickable" nitrilotriacetic acid and iminodiacetic acid tags.
- Authors: Loh CT, Graham B, Abdelkader EH, Tuck KL, Otting G
- Issue date: 2015 Mar 23
- A protocol for the refinement of NMR structures using simultaneously pseudocontact shift restraints from multiple lanthanide ions.
- Authors: Sala D, Giachetti A, Luchinat C, Rosato A
- Issue date: 2016 Nov
- NMR structure determination of protein-ligand complexes by lanthanide labeling.
- Authors: Pintacuda G, John M, Su XC, Otting G
- Issue date: 2007 Mar
- Pseudocontact Shift-Driven Iterative Resampling for 3D Structure Determinations of Large Proteins.
- Authors: Pilla KB, Otting G, Huber T
- Issue date: 2016 Jan 29