Determination of Relative N−HN, N−C‘, Cα−C‘, and Cα−Hα Effective Bond Lengths in a Protein by NMR in a Dilute Liquid Crystalline Phase

Ottiger, Marcel; Bax, Ad

doi:10.1021/ja9826791

Cited by 222 publications

(216 citation statements)

References 63 publications

(130 reference statements)

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“…In addition, order parameters and internuclear distances for atom pairs were set as described in reference. 39 HetNOEs were measured with an 15 N-[ 1 H]-NOE-TROSY experiment. The errors result from peak volume fitting inaccuracies.…”

Section: Methodsmentioning

confidence: 99%

Conformational stability and activity of p73 require a second helix in the tetramerization domain

et al. 2009

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p73 and p63, the two ancestral members of the p53 family, are involved in neurogenesis, epithelial stem cell maintenance and quality control of female germ cells. The highly conserved oligomerization domain (OD) of tumor suppressor p53 is essential for its biological functions, and its structure was believed to be the prototype for all three proteins. However, we report that the ODs of p73 and p63 differ from the OD of p53 by containing an additional a-helix that is not present in the structure of the p53 OD. Deletion of this helix causes a dissociation of the OD into dimers; it also causes conformational instability and reduces the transcriptional activity of p73. Moreover, we show that ODs of p73 and p63 strongly interact and that a large number of different heterotetramers are supported by the additional helix. Detailed analysis shows that the heterotetramer consisting of two homodimers is thermodynamically more stable than the two homotetramers. No heterooligomerization between p53 and the p73/p63 subfamily was observed, supporting the notion of functional orthogonality within the p53 family. p53, a well-known tumor suppressor that is mutated in more than 50% of all human tumors, induces genes leading either to cell-cycle arrest or to apoptosis. The discovery of two proteins with a high sequence identity to p53, called p63 and p73, has sparked speculations that tumor suppression is carried out by the combined action of several members of this protein family, for example, by direct interaction through heterooligomerization. Knockout mouse studies with p63 and p73, the two ancestral members, 1 have shown functional roles distinct from p53: p63 is essential for maintaining epithelial stem cells 2,3 and for protecting the genomic stability of oocytes, 4 whereas p73 is involved in neurogenesis, sensory pathways and homeostatic control. 5 p73 is further known as an important inducer of apoptosis in response to DNA damage. 6 Although only few mutations of p63 and p73 have been found in human tumors so far, overexpression of p63 is often observed in squamous cell carcinoma, 7-10 which has been shown to suppress p73-dependent apoptosis. 11 Among all p53 family members, including those from invertebrate species, the DNA binding domain is the most conserved domain, 12-15 followed by the oligomerization domain (OD), which is indispensable for the biological function of all p53 protein family members. [16][17][18] It is a structural domain that forms a tetramer, and mutations within the OD that inhibit tetramerization of p53 result in greatly reduced transcriptional activity. 19 In addition, several protein-protein interactions and posttranslational modifications require the tetrameric state as well, 16 and mutations in the OD of p53 that prevent oligomerization have been identified in human cancers. 20,21 Owing to its functional importance, the OD of p53 has been the target of several structure determination projects. 22,23 The p53 tetramer consists of a dimer of dimers, with each monomer contributing one b-strand and o...

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Section: Methodsmentioning

confidence: 99%

Conformational stability and activity of p73 require a second helix in the tetramerization domain

et al. 2009

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“…The use of this model requires an assumed dipole-dipole coupling constant or the carbon-proton distance. We choose here to use r CH =111.7 pm, based on the work of Ottiger and Bax (Ottiger and Bax 1998). Making use of Eqs.…”

Section: T Although Partial Spectral Overlap May Occur For Residuementioning

confidence: 99%

Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation

2013

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“…29 The reduced spectral density function of the particular Larmor frequencies, ω, for an isotropically tumbling molecule with limited and rapid internal flexibility can be approximated by:…”

Section: Introductionmentioning

confidence: 99%

Conformation and Dynamics at a Flexible Glycosidic Linkage Revealed by NMR Spectroscopy and Molecular Dynamics Simulations: Analysis of β-l-Fucp-(1→6)-α-d-Glcp-OMe in Water Solution

2013

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The intrinsic flexibility of carbohydrates facilitates different three-dimensional structures in response to altered environments. At glycosidic (1→6)-linkages three torsion angles are variable and herein the conformation and dynamics of β-L-Fucp-(1→6)-α-D-Glcp-OMe are investigated using a combination of NMR spectroscopy and molecular dynamics (MD) simulations. The disaccharide shows evidence of conformational averaging for the ψ and ω torsion angles, best explained by a four-state conformational distribution. Notably, there is a significant population of conformations having ψ = 85° (clinal) in addition to those having ψ = 180° (antiperiplanar). Moderate differences in13 C R 1 relaxation rates are found to be best explained by axially symmetric tumbling in combination with minor differences in librational motion for the two residues, whereas the isomerization motions are occurring too slowly to be contributing significantly to the observed relaxation rates. The MD simulation was found to give a reasonably good agreement with experiment, especially with respect to diffusive properties among which the rotational anisotropy,, is found to be 2.35. The force field employed showed too narrow ω torsion angles in the gauche-trans and gauche-gauche states as well as overestimating the population of the gauchetrans conformer. This information can subsequently be used in directing parameter developments and emphasizes the need for refinement of force fields for (1→6)-linked carbohydrates.

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Determination of Relative N−H^N, N−C‘, C^α−C‘, and C^α−H^α Effective Bond Lengths in a Protein by NMR in a Dilute Liquid Crystalline Phase

Cited by 222 publications

References 63 publications

Conformational stability and activity of p73 require a second helix in the tetramerization domain

Conformational stability and activity of p73 require a second helix in the tetramerization domain

Dynamics of exocyclic groups in the Escherichia coli O91 O-antigen polysaccharide in solution studied by carbon-13 NMR relaxation

Conformation and Dynamics at a Flexible Glycosidic Linkage Revealed by NMR Spectroscopy and Molecular Dynamics Simulations: Analysis of β-l-Fucp-(1→6)-α-d-Glcp-OMe in Water Solution

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