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. 2013 Oct 15;110(42):16826-31.
doi: 10.1073/pnas.1315525110. Epub 2013 Sep 30.

Physics-based method to validate and repair flaws in protein structures

Osvaldo A Martin  1 , Yelena A Arnautova, Alejandro A Icazatti, Harold A Scheraga, Jorge A Vila
Affiliations

Physics-based method to validate and repair flaws in protein structures

Osvaldo A Martin et al. Proc Natl Acad Sci U S A. .

Abstract

A method that makes use of information provided by the combination of (13)C(α) and (13)C(β) chemical shifts, computed at the density functional level of theory, enables one to (i) validate, at the residue level, conformations of proteins and detect backbone or side-chain flaws by taking into account an ensemble average of chemical shifts over all of the conformations used to represent a protein, with a sensitivity of ∼90%; and (ii) provide a set of (χ1/χ2) torsional angles that leads to optimal agreement between the observed and computed (13)C(α) and (13)C(β) chemical shifts. The method has been incorporated into the CheShift-2 protein validation Web server. To test the reliability of the provided set of (χ1/χ2) torsional angles, the side chains of all reported conformations of five NMR-determined protein models were refined by a simple routine, without using NOE-based distance restraints. The refinement of each of these five proteins leads to optimal agreement between the observed and computed (13)C(α) and (13)C(β) chemical shifts for ∼94% of the flaws, on average, without introducing a significantly large number of violations of the NOE-based distance restraints for a distance range ≤ 0.5 , in which the largest number of distance violations occurs. The results of this work suggest that use of the provided set of (χ1/χ2) torsional angles together with other observables, such as NOEs, should lead to a fast and accurate refinement of the side-chain conformations of protein models.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Structures of ubiquitin in the left-hand column were determined by NMR spectroscopy; those in the right-hand column were determined by X-ray crystallography. Structures in A and B pertain to validation of proteins 1D3Z and 1UBQ, respectively, by using only 13Cα chemical shifts; structures in C and D pertain to validation of proteins 1D3Z and 1UBQ, respectively, by using only 13Cβ chemical shifts; and structures in E and F pertain to validation of proteins 1D3Z and 1UBQ, respectively, by using a combination of 13Cα and 13Cβ chemical shifts. All of the residues for which the agreement between observed and computed 13Cα or 13Cβ chemical shifts (A–D) can be improved, i.e., by varying the side-chain torsional angles to any of the solutions provided by the CheShift-2 Web server, are highlighted in blue in E and F. Those residues showing good, marginally good, and poor agreement with the observed chemical shift values are highlighted in green, yellow, and red, respectively.
Fig. 2.
Fig. 2.
(A) Graphic validation, in terms of both the 13Cα and 13Cβ chemical shifts, for the 10 models of ubiquitin 1D3Z in which 39 residues are highlighted in blue to indicate that their validation could be improved by varying their side-chain torsional angles by using the CheShift-2 Web server solutions (SI Appendix, Table S2). (B) The refined 10 models obtained by using the CheShift-2 Web server solutions. A total of 34 residues, now colored in green, were improved, while the remaining five residues are still colored in blue because any of the solutions provided by the CheShift-2 Web server leads to an unacceptably large atomic overlapping.
Fig. 3.
Fig. 3.
(A) Black- and gray-filled bars denote the average number, per conformer, of NOE-derived distance restraint violations obtained from the original and the refined ensembles of 10 conformations of 1D3Z, respectively. The violations are grouped within intervals of 0.5 Ǻ. At a given interval, e.g., 1.0 Ǻ, the heights of the bars represent the accumulated number of violations (X), which are in the range 0.5 < X ≤ 1.0 Ǻ. (B) The difference, as gray bars, between the total number of NOE-derived distance restraint violations after and before the refinement as a function of the conformation number for each of the 10 conformations of 1D3Z. The black-filled square for each conformation represent the total number of repaired flaws.
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