Validation of Protein Structure from Anisotropic Carbonyl Chemical Shifts in a Dilute Liquid Crystalline Phase

Cornilescu, Gabriel; Marquardt, John L.; Ottiger, Marcel; Bax, Ad

doi:10.1021/ja9812610

Cited by 872 publications

(1,052 citation statements)

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“…For the individual ubiquitin (Ub) domains in di-ubiquitin (Ub 2 ) we used NMR solution structure of monomeric Ub determined by Cornilescu at al. 35 (PDB code 1D3Z, where we clipped the flexible C-terminus beyond Arg72). As shown earlier12,14 the backbone structure of each Ub domain in Ub 2 is practically identical to that of an isolated Ub.…”

Section: Methodsmentioning

confidence: 99%

Structural Assembly of Multidomain Proteins and Protein Complexes Guided by the Overall Rotational Diffusion Tensor

Ryabov

Fushman

2007

J. Am. Chem. Soc.

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We present a simple and robust approach that uses the overall rotational diffusion tensor as a structural constraint for domain positioning in multidomain proteins and protein-protein complexes. This method offers the possibility to use NMR relaxation data for detailed structure characterization of such systems provided the structures of individual domains are available. The proposed approach extends the concept of using long-range information contained in the overall rotational diffusion tensor. In contrast to the existing approaches, we use both the principal axes and principal values of protein's rotational diffusion tensor to determine not only the orientation but also the relative positioning of the individual domains in a protein. This is achieved by finding the domain arrangement in a molecule that provides the best possible agreement with all components of the overall rotational diffusion tensor derived from experimental data. The accuracy of the proposed approach is demonstrated for two protein systems with known domain arrangement and parameters of the overall tumbling: the HIV-1 protease homodimer and Maltose Binding Protein. The accuracy of the method and its sensitivity to domain positioning is also tested using computer-generated data for three protein complexes, for which the experimental diffusion tensors are not available. In addition, the proposed method is applied here to determine, for the first time, the structure of both open and closed conformations of Lys48-linked di-ubiquitin chain, where domain motions render impossible accurate structure determination by other methods. The proposed method opens new avenues for improving structure characterization of proteins in solution.

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Section: Methodsmentioning

confidence: 99%

Structural Assembly of Multidomain Proteins and Protein Complexes Guided by the Overall Rotational Diffusion Tensor

Ryabov

Fushman

2007

J. Am. Chem. Soc.

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“…J DD (ω) =J CSA (ω)=J(ω). In general, however 62,31 , J DD (ω) ≠ J CSA (ω), and a correction for the difference between the spectral densities can be included as: (22) where f is the correction factor: f = [J DD (0)/J CSA (0) First, the very nature of the chemical shielding suggests that it should fluctuate when local environment of a nucleus changes as a result of internal motions in a protein.…”

Section: Possible Sources Of Systematic Errors In 15 N Csa Determinatmentioning

confidence: 99%

Variability of the ¹⁵N Chemical Shielding Tensors in the B3 Domain of Protein G from ¹⁵N Relaxation Measurements at Several Fields. Implications for Backbone Order Parameters

2006

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We applied a combination of 15 N relaxation and CSA/dipolar cross-correlation measurements at five magnetic fields (9.4, 11.7, 14.1, 16.4, and 18.8 Tesla) to determine the 15 N chemical shielding tensors for backbone amides in protein G in solution. The data were analyzed using various modelindependent approaches and those based on Lipari-Szabo approximation, all of them yielding similar results. The results indicate a range of site-specific values of the anisotropy (CSA) and orientation of the 15 N chemical shielding tensor, similar to those in ubiquitin. Assuming a Gaussian distribution of the 15 N CSA values, the mean anisotropy is -173.9 to -177.2 ppm (for 1.02-Å NH-bond length) and the site-so-site CSA variability is ±17.6 to ±21.4 ppm, depending on the method used. This CSA variability is significantly larger than derived previously for ribonuclease H or recently, using "metaanalysis" for ubiquitin. Standard interpretation of 15 N relaxation studies of backbone dynamics in proteins involves an a priori assumption of a uniform 15 N CSA. We show that this assumption leads to a significant discrepancy between the order parameters obtained at different fields. Using the sitespecific CSAs obtained from our study removes this discrepancy and allows simultaneous fit of relaxation data at all five fields to Lipari-Szabo spectral densities. These findings emphasize the necessity of taking into account the variability of 15 N CSA for accurate analysis of protein dynamics from 15 N relaxation measurements.

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“…The reference frame employed for projecting the simulated trajectories was built through principal components analysis of the covariance matrix of Cα positions for residues 2−70 as retrieved from 72 high-resolution X-ray structures of human ubiquitin aligned to model 1 of PDB ID 1D3Z. 81 Projection of the trajectories on the reference frame was also preceded by alignment to this reference structure. Logarithmic probability distributions were built by projecting the trajectories on a reference frame, binning them into a grid and counting the number of frames inside each cell of the grid (N i ) to compute −k B T ln(N i /N 0 ), where N 0 corresponds to the most populated bin thus setting the offset.…”

Section: ■ Introductionmentioning

confidence: 99%

Dissecting the Effects of Concentrated Carbohydrate Solutions on Protein Diffusion, Hydration, and Internal Dynamics

et al. 2014

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We present herein a thorough description of the effects of high glucose concentrations on the diffusion, hydration and internal dynamics of ubiquitin, as predicted from extensive molecular dynamics simulations on several systems described at fully atomistic level. We observe that the protein acts as a seed that speeds up the natural propensity of glucose to cluster at high concentration; the sugar molecules thus aggregate around the protein trapping it inside a dynamic cage. This process extensively dehydrates the protein surface, restricts the motions of the remaining water molecules, and drags the large-scale, collective motions of protein atoms slowing down the rate of exploration of the conformational space despite only a slight dampening of fast, local dynamics. We discuss how these effects could be relevant to the function of sugars as preservation agents in biological materials, and how crowding by small sticky molecules could modulate proteins across different reaction coordinates inside the cellular cytosol. ■ INTRODUCTIONSugars play roles as agents for the preservation of biological material in nature and in biotechnological manufacture, 1−3 most importantly through their capacity to stabilize proteins against cold and hot denaturation, both in solution and in the solid state. 4−7 Other potential effects of sugars on protein properties have been less explored, but changes in activity, dynamics, and regulation can be expected by analogy to the effects known to be caused by high concentrations of other molecules. Whereas most hydrophilic molecules have the capacity to agglomerate and disrupt water structure, sugars and polyols generally have exceptionally large solubilities, which allows them to strongly dehydrate other molecules and to cluster at very high concentrations forming glassy states. 8−11 Because of these special properties, questions about the effects of sugars on protein properties are intimately related to those revolving around the effects of viscosity, molecular crowding, encapsulation and even vitrification on proteins, all meeting at the crossroads between chemistry, biology, medicine, and applications in the food and pharmacological industries. Within all these closely related fields, the effects of high concentrations of small hydrophilic molecules on protein stability and translational and rotational diffusion have been explored, but studies of their effects on protein hydration and on internal protein dynamics are scarce. 4−7,12−16 More specifically to sugars, reports on the structure and dynamics of sugar-only solutions abound, 8,17−23 but works dealing with proteins in sugar solutions are mostly focused on the stability of the proteins and do not pay much attention to other effects related to hydration, diffusion, and internal mobility. 4−7 We recently reported in a preliminary work that concentrated glucose solutions can perturb protein dynamics by restricting exploration of the conformational space, through a mechanism that presumably involves protein−sugar interac...

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Validation of Protein Structure from Anisotropic Carbonyl Chemical Shifts in a Dilute Liquid Crystalline Phase

Abstract: Q ) rms(∆δ meas -∆δ pred )/rms(∆δ meas )(3) Communications to the Editor

Cited by 872 publications

References 14 publications

Structural Assembly of Multidomain Proteins and Protein Complexes Guided by the Overall Rotational Diffusion Tensor

Structural Assembly of Multidomain Proteins and Protein Complexes Guided by the Overall Rotational Diffusion Tensor

Variability of the ¹⁵N Chemical Shielding Tensors in the B3 Domain of Protein G from ¹⁵N Relaxation Measurements at Several Fields. Implications for Backbone Order Parameters

Dissecting the Effects of Concentrated Carbohydrate Solutions on Protein Diffusion, Hydration, and Internal Dynamics

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Validation of Protein Structure from Anisotropic Carbonyl Chemical Shifts in a Dilute Liquid Crystalline Phase

Abstract: Q ) rms(∆δ meas -∆δ pred )/rms(∆δ meas )(3) Communications to the Editor

Cited by 872 publications

References 14 publications

Structural Assembly of Multidomain Proteins and Protein Complexes Guided by the Overall Rotational Diffusion Tensor

Structural Assembly of Multidomain Proteins and Protein Complexes Guided by the Overall Rotational Diffusion Tensor

Variability of the 15N Chemical Shielding Tensors in the B3 Domain of Protein G from 15N Relaxation Measurements at Several Fields. Implications for Backbone Order Parameters

Dissecting the Effects of Concentrated Carbohydrate Solutions on Protein Diffusion, Hydration, and Internal Dynamics

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Variability of the ¹⁵N Chemical Shielding Tensors in the B3 Domain of Protein G from ¹⁵N Relaxation Measurements at Several Fields. Implications for Backbone Order Parameters