Liquid-state NMR spectroscopy is a powerful technique to elucidate binding properties of ligands on proteins. Ligands binding in hydrophobic pockets are often in close proximity to methyl groups and binding can lead to subtle displacements of methyl containing side chains to accommodate the ligand. To establish whether pseudocontact shifts can be used to characterize ligand binding and the effects on methyl groups, the N-terminal domain of HSP90 was tagged with caged lanthanoid NMR probe 5Â*at three positions and titrated with a ligand. Binding was monitored using the resonances of leucine and valine methyl groups. The pseudocontact shifts (PCS) caused by ytterbium result in enhanced dispersion of the methyl spectrum, allowing more resonances to be observed. The effects of tag attachment on the spectrum and ligand binding are small. Significant changes in PCS were observed upon ligand binding, indicating displacements of several methyl groups. By determining the cross-section of PCS iso-surfaces generated by two or three paramagnetic centers, the new position of a methyl group can be estimated, showing displacements in the range of 1â??3Â*Ã? for methyl groups in the binding site. The information about such subtle but significant changes may be used to improve docking studies and can find application in fragment-based drug discovery.
Complete assignment of Ala, Ile, Leu, Met and Val methyl groups of human blood group A and B glycosyltransferases using lanthanide-induced pseudocontact shifts and methylâ??methyl NOESY
Complete assignment of Ala, Ile, Leu, Met and Val methyl groups of human blood group A and B glycosyltransferases using lanthanide-induced pseudocontact shifts and methylâ??methyl NOESY
Abstract
Human blood group A and B glycosyltransferases (GTA, GTB) are highly homologous glycosyltransferases. A number of high-resolution crystal structures is available showing that these enzymes convert from an open conformation into a catalytically active closed conformation upon substrate binding. However, the mechanism of glycosyltransfer is still under debate,...
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04-26-2018 04:36 PM
Methyl group assignment using pseudocontact shifts with PARAssign
Methyl group assignment using pseudocontact shifts with PARAssign
Abstract
A new version of the program PARAssign has been evaluated for assignment of NMR resonances of the 76 methyl groups in leucines, isoleucines and valines in a 25Â*kDa protein, using only the structure of the protein and pseudocontact shifts (PCS) generated with a lanthanoid tag at up to three attachment sites. The number of reliable assignments depends strongly on two factors. The principle axes of the magnetic susceptibility tensors of the paramagnetic centers should not be...
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11-27-2017 01:29 PM
[NMR paper] Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange.
Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange.
Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange.
Angew Chem Int Ed Engl. 2017 Aug 28;:
Authors: Gao J, Liang E, Ma R, Li F, Liu Y, Liu J, Jiang L, Li C, Dai H, Wu J, Su X, He W, Ruan K
Abstract
The characterization of protein-ligand interaction modes becomes recalcitrant in the NMR intermediate exchange regime as the...
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08-29-2017 05:35 PM
[NMR paper] Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange
Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange
The characterization of protein-ligand interaction modes becomes recalcitrant in the NMR intermediate exchange regime as the interface resonances are broadened beyond detection. Here, we determined the 19F low-populated bound-state pseudocontact shifts (PCSs) of mono- and di-fluorinated inhibitors of the BRM bromodomain using a highly-skewed protein/ligand ratio. The bound-state 19F PCSs were retrieved from 19F chemical exchange saturation transfer (CEST) in...
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08-29-2017 01:33 AM
Sequence-specific assignment of methyl groups from the neuronal SNARE complex using lanthanide-induced pseudocontact shifts
Sequence-specific assignment of methyl groups from the neuronal SNARE complex using lanthanide-induced pseudocontact shifts
Abstract
Neurotransmitter release depends critically on the neuronal SNARE complex formed by syntaxin-1, SNAP-25 and synaptobrevin, as well as on other proteins such as Munc18-1, Munc13-1 and synaptotagmin-1. Although three-dimensional structures are available for these components, it is still unclear how they are assembled between the synaptic vesicle and plasma membranes to trigger fast, Ca2+-dependent membrane fusion. Methyl...
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12-18-2016 05:04 AM
An encodable lanthanide binding tag with reduced size and flexibility for measuring residual dipolar couplings and pseudocontact shifts in large proteins
An encodable lanthanide binding tag with reduced size and flexibility for measuring residual dipolar couplings and pseudocontact shifts in large proteins
Abstract
Metal ions serve important roles in structural biology applications from long-range perturbations seen in magnetic resonance experiments to electron-dense signatures in X-ray crystallography data; however, the metal ion must be secured in a molecular framework to achieve the maximum benefit. Polypeptide-based lanthanide-binding tags (LBTs) represent one option that can be directly encoded...
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01-04-2016 07:49 PM
Pulse EPR-enabled interpretation of scarce pseudocontact shifts induced by lanthanide binding tags
Pulse EPR-enabled interpretation of scarce pseudocontact shifts induced by lanthanide binding tags
Abstract
Pseudocontact shifts (PCS) induced by tags loaded with paramagnetic lanthanide ions provide powerful long-range structure information, provided the location of the metal ion relative to the target protein is known. Usually, the metal position is determined by fitting the magnetic susceptibility anisotropy (Î?Ï?) tensor to the 3D structure of the protein in an 8-parameter fit, which requires a large set of PCSs to be reliable. In an alternative...
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11-23-2015 06:58 PM
Generation of Pseudocontact Shifts in Protein NMR Spectra with a Genetically Encoded Cobalt(II)-Binding Amino Acid.
Generation of Pseudocontact Shifts in Protein NMR Spectra with a Genetically Encoded Cobalt(II)-Binding Amino Acid.
Generation of Pseudocontact Shifts in Protein NMR Spectra with a Genetically Encoded Cobalt(II)-Binding Amino Acid.
Angew Chem Int Ed Engl. 2011 Jan 17;50(3):692-4
Authors: Nguyen TH, Ozawa K, Stanton-Cook M, Barrow R, Huber T, Otting G
Methyl group reorientation under ligand binding probed by pseudocontact shifts
Linear Mode
