RDC‐assisted modeling of symmetric protein homo‐oligomers

Wang, Xu; Bansal, Sonal; Jiang, Mei; Prestegard, James H.

doi:10.1110/ps.073395108

Cited by 36 publications

(68 citation statements)

References 48 publications

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“…10,11 RDCs collected on 1 H-15 N pairs in structured regions of the protein can be used to define the alignment frames in multiple media, and for a tight dimer the symmetry axis can be identified as the alignment frame axis which is preserved among different alignment media. 5,34 For weak dimers it is important to realize that the measured RDC values will be the average of the monomer and dimer values. For SeR13 this complication could be addressed by extrapolating RDCs measured at a series of concentrations to RDCs at infinite concentration.…”

Section: Symmetry Axis Constraints On Ser13 Dimerizationmentioning

confidence: 99%

“…5 Details of this procedure are given in the methods section. Out of 5107 possible dimeric models (each with different X and Z grid points), 29 models gave both acceptable RDC correlations and residue pair scores.…”

Section: Constructing the Dimeric Model Of Ser13mentioning

confidence: 99%

“…We have shown previously that residual dipolar coupling (RDC) data can be used to characterize tightly associated homodimeric structures. 5 Here we present an NMR methodology that can provide structures of even weakly associating homo-oligomer complexes in the absence of NOE data.…”

Section: Introductionmentioning

confidence: 99%

“…10,11 The NOE independent approach described here follows previous work of Wang et al who used a combination of restraints from RDCs and a residue pair scoring function to generate a dimer model of a high affinity complex. 5 The work built on an X-ray crystallography study of the YkuJ protein (Northeast Structural Genomics target SR360) which provided a reliable monomer structure but two alternative dimer structures. Two independent alignment media were used to remove the three-fold ambiguity in identifying the symmetry axis.…”

Section: Introductionmentioning

confidence: 99%

“…5,13,14 A weakly associated hetero-oligomeric complex was solved by Ortega-Roldan et al using RDC orientational restraints along with chemical shift perturbations. 13 Dimer RDCs were extracted by titrating isotopically-enriched monomer A with unlabeled monomer B and the complex was assembled using the alignment tensors calculated from four types of RDC measured in a single medium.…”

Section: Introductionmentioning

confidence: 99%

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Three‐dimensional structure of the weakly associated protein homodimer SeR13 using RDCs and paramagnetic surface mapping

et al. 2010

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The traditional NMR-based method for determining oligomeric protein structure usually involves distinguishing and assigning intra-and intersubunit NOEs. This task becomes challenging when determining symmetric homo-dimer structures because NOE cross-peaks from a given pair of protons occur at the same position whether intra-or intersubunit in origin. While there are isotope-filtering strategies for distinguishing intra from intermolecular NOE interactions in these cases, they are laborious and often prove ineffectual in cases of weak dimers, where observation of intermolecular NOEs is rare. Here, we present an efficient procedure for weak dimer structure determination based on residual dipolar couplings (RDCs), chemical shift changes upon dilution, and paramagnetic surface perturbations. This procedure is applied to the Northeast Structural Genomics Consortium protein target, SeR13, a negatively charged Staphylococcus epidermidis dimeric protein (K d 3.4 6 1.4 mM) composed of 86 amino acids. A structure determination for the monomeric form using traditional NMR methods is presented, followed by a dimer structure determination using docking under orientation constraints from RDCs data, and scoring under residue pair potentials and shape-based predictions of RDCs. Validation using paramagnetic surface perturbation and chemical shift perturbation data acquired on sample dilution is also presented. The general utility of the dimer structure determination procedure and the possible relevance of SeR13 dimer formation are discussed.

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Section: Symmetry Axis Constraints On Ser13 Dimerizationmentioning

confidence: 99%

Section: Constructing the Dimeric Model Of Ser13mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Three‐dimensional structure of the weakly associated protein homodimer SeR13 using RDCs and paramagnetic surface mapping

et al. 2010

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Structural and biophysical characterization of the type VII collagen vWFA2 subdomain leads to identification of two binding sites

et al. 2020

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Type VII collagen is an extracellular matrix protein, which is important for skin stability; however, detailed information at the molecular level is scarce. The second vWFA (von Willebrand factor type A) domain of type VII collagen mediates important interactions, and immunization of mice induces skin blistering in certain strains. To understand vWFA2 function and the pathophysiological mechanisms leading to skin blistering, we structurally characterized this domain by X‐ray crystallography and NMR spectroscopy. Cell adhesion assays identified two new interactions: one with β1 integrin via its RGD motif and one with laminin‐332. The latter interaction was confirmed by surface plasmon resonance with a KD of about 1 mm. These data show that vWFA2 has additional functions in the extracellular matrix besides interacting with type I collagen.

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