Efficient and accurate determination of the overall rotational diffusion tensor of a molecule from 15N relaxation data using computer program ROTDIF

Walker, Olivier; Varadan, Ranjani; Fushman, David

doi:10.1016/j.jmr.2004.03.019

Cited by 79 publications

(130 citation statements)

References 28 publications

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“…In contrast, the backbone order parameters in the rest of the UQ1-UBA construct (S 2 = 0.84 ± 0.04 for residues 546-584 and 0.86 ± 0.04 in the elements of secondary structure) are at the level typical for a well-folded protein, thus confirming that the structure of UQ1-UBA is well defined. Analysis using ROTDIF 45 program showed that the UQ1-UBA structure obtained here is in good agreement with the observed orientational dependence of the 15 N relaxation rates. The overall rotational correlation time (Table 3) obtained from this analysis (3.30 ± 0.13 ns) agrees well with the expected tumbling time of UQ1-UBA (3.56 ns predicted using HYDRONMR 46 ), thus confirming that UQ1-UBA behaves as a monomer in solution under these conditions.…”

Section: Structure Of the Uq1-uba/monoub Complexsupporting

confidence: 85%

“…Interestingly, in Ub-bound UQ1-UBA, the 15 N R 2 / R 1 ratio (which is sensitive to NH bond's orientation relative to the global rotational diffusion tensor 45 ) is systematically higher in helix α1 (11.2 ± 0.7) than in helices α2 and α3 (8.4 ± 0.4 and 7.9 ± 0.4, respectively). This observation supports the calculated structure of the UQ1-UBA complex (Figs.…”

Section: Structure Of the Uq1-uba/monoub Complexmentioning

confidence: 98%

“…The experiments were carried out on UQ1-UBA alone as well as on both UQ1-UBA and Ub in the complex; the experimental data were analyzed as detailed elsewhere. 47 Briefly, the 15 N relaxation rates and heteronuclear NOEs were analyzed using ROTDIF 45 and assuming various overall rotational diffusion models (Supplementary Data), and the resulting diffusion tensor was used as input into computer program DYNAMICS 47,65 for extracting the "model-free" parameters characterizing backbone dynamics.…”

Section: N Relaxation Measurements and Analysis Of Protein Dynamicsmentioning

confidence: 99%

“…Characteristics of the overall rotational diffusion derived from 15 N relaxation data for Ub and UQ1-UBA, free in solution and in the complex, using ROTDIF analysis45 …”

mentioning

confidence: 99%

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Affinity Makes the Difference: Nonselective Interaction of the UBA Domain of Ubiquilin-1 with Monomeric Ubiquitin and Polyubiquitin Chains

Zhang

Raasi

Fushman

2008

Journal of Molecular Biology

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147

184

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Ubiquilin/PLIC proteins belong to the family of UBL-UBA proteins implicated in the regulation of the ubiquitin-dependent proteasomal degradation of cellular proteins. A human presenilin-interacting protein, ubiquilin-1, has been suggested as potential therapeutic target for treating Huntington's disease. Ubiquilin's interactions with mono- and polyubiquitins are mediated by its UBA domain, which is one of the tightest ubiquitin binders among known ubiquitin-binding domains. Here we report the three-dimensional structure of the UBA domain of ubiquilin-1 (UQ1-UBA) free in solution and in complex with ubiquitin. UQ1-UBA forms a compact three-helix bundle structurally similar to other known UBAs, and binds to the hydrophobic patch on ubiquitin with a K(d) of 20 microM. To gain structural insights into UQ1-UBA's interactions with polyubiquitin chains, we have mapped the binding interface between UQ1-UBA and Lys48- and Lys63-linked di-ubiquitins and characterized the strength of UQ1-UBA binding to these chains. Our NMR data show that UQ1-UBA interacts with the individual ubiquitin units in both chains in a mode similar to its interaction with mono-ubiquitin, although with an improved binding affinity for the chains. Our results indicate that, in contrast to UBA2 of hHR23A that has strong binding preference for Lys48-linked chains, UQ1-UBA shows little or no binding selectivity toward a particular chain linkage or between the two ubiquitin moieties in the same chain. The structural data obtained in this study provide insights into the possible structural reasons for the diversity of polyubiquitin chain recognition by UBA domains.

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Section: Structure Of the Uq1-uba/monoub Complexsupporting

confidence: 85%

Section: Structure Of the Uq1-uba/monoub Complexmentioning

confidence: 98%

Section: N Relaxation Measurements and Analysis Of Protein Dynamicsmentioning

confidence: 99%

“…Characteristics of the overall rotational diffusion derived from 15 N relaxation data for Ub and UQ1-UBA, free in solution and in the complex, using ROTDIF analysis45 …”

mentioning

confidence: 99%

See 2 more Smart Citations

Affinity Makes the Difference: Nonselective Interaction of the UBA Domain of Ubiquilin-1 with Monomeric Ubiquitin and Polyubiquitin Chains

Zhang

Raasi

Fushman

2008

Journal of Molecular Biology

Self Cite

147

184

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“…Methods have been proposed to determine this anisotropy using heteronuclear [246] or homonuclear [247] relaxation rates, in the absence of any structural information. If the structure of the observed molecule is know, the anisotropy of diffusion can be determined from the relaxation data by simulated annealing [248,249], by optimization [250], by a Bayesian approach [251] or by a graphical method [252].…”

Section: Describing the Relaxation Measurements In Terms Of Internal mentioning

confidence: 99%

Quantitative Analysis of Biomolecular NMR Spectra: A Prerequisite for the Determination of the Structure and Dynamics of Biomolecules

Malliavin

2006

COC

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Nuclear Magnetic Resonance (NMR) became during the two last decades an important method for biomolecular structure determination. NMR permits to study biomolecules in solution and gives access to the molecular flexibility at atomic level on a complete structure: in that respect, it is occupying a unique place i n structural biology. During the first years of its development, NMR was trying to meet the requirements previously defined in X-ray crystallography. But, NMR then started to determine its own criteria for the definition of a structure. Indeed, the atomic coordinates of an NMR structure are calculated using restraints on geometrical parameters (angles and distances) of the structure, which are only indirectly related to atom positions: in that respect, NMR and X-ray crystallography are very different. The indirect relation between the NMR measurements and the molecular structure and dynamics makes critical the precision and the interpretation of the NMR parameters and the development of quantitative analysis methods. The methods published since 1997 for liquid-NMR of proteins are reviewed here. First, methods for structure determination are presented, as well as methods for spectral assignment and for structure quality assessment. Second, the quantitative analysis of structure mobility i s reviewed.

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Identification of primary and secondary UBA footprints on the surface of ubiquitin in cell‐mimicking crowded solution

et al. 2017

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Despite significant advancements in our understanding of ubiquitin-mediated signaling, the influence of the intracellular environment on formation of transient ubiquitin-partner complexes remains poorly explored. In our work, we introduce macromolecular crowding as a first level of complexity towards the imitation of a cellular environment in the study of such interactions. Using NMR spectroscopy, we find that the stereospecific complex of ubiquitin and ubiquitin-associated domain (UBA) is minimally perturbed by the crowding agent Ficoll. However, in addition to the primary canonical recognition patch on ubiquitin, secondary patches are identified, indicating that in cell-mimicking crowded solution, UBA contacts ubiquitin at multiple sites.

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Efficient and accurate determination of the overall rotational diffusion tensor of a molecule from 15N relaxation data using computer program ROTDIF

Cited by 79 publications

References 28 publications

Affinity Makes the Difference: Nonselective Interaction of the UBA Domain of Ubiquilin-1 with Monomeric Ubiquitin and Polyubiquitin Chains

Affinity Makes the Difference: Nonselective Interaction of the UBA Domain of Ubiquilin-1 with Monomeric Ubiquitin and Polyubiquitin Chains

Quantitative Analysis of Biomolecular NMR Spectra: A Prerequisite for the Determination of the Structure and Dynamics of Biomolecules

Identification of primary and secondary UBA footprints on the surface of ubiquitin in cell‐mimicking crowded solution

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