Free energy calculations on the stability of the 14-3-3ζ protein

Jandová, Zuzana; Trošanová, Zuzana; Weisová, Veronika; Oostenbrink, Chris; Hritz, Jozef

doi:10.1016/j.bbapap.2017.11.012

Cited by 9 publications

(12 citation statements)

References 49 publications

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“…For the remaining cases, fSASA was used to categorize the mutation site in the input structure as buried (fSASA < 10%; 44 total cases) and solvent exposed (fSASA ≥ 10%; 106 total cases). The definition of fSASA is taken to be the fraction of the maximum possible solvent accessible surface area in a tripeptide configuration experienced by the residue in the interface [13]. We note here that all mutations with a significant favorable effect on binding are contained in the latter set, and FEP performance is good (RMSE 1.23) on the set of non-buried positions.…”

Section: Models and Methodsmentioning

confidence: 99%

“…[11] consisted exclusively of mutations that did not change the net charge on the system; that is, charge changing mutations such as Ala to Glu were excluded from the data set. Although some attempts at applying explicit solvent free energy calculations to problems such as protein folding exist [13], we deliberately set aside mutations in the latter category due to the well-known technical difficulties in performing alchemical simulations in which the net charge on the system is altered [14], [15], [16], [17], [18], [19]. We apply a variant of the co-alchemical water FEP approach first proposed by Wallace and Shen [15] and Chen et al .…”

Section: Introductionmentioning

confidence: 99%

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Relative Binding Affinity Prediction of Charge-Changing Sequence Mutations with FEP in Protein–Protein Interfaces

Clark

Negron

Hauser

et al. 2019

Journal of Molecular Biology

149

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Building on the substantial progress that has been made in using free energy perturbation (FEP) methods to predict the relative binding affinities of small molecule ligands to proteins, we have previously shown that results of similar quality can be obtained in predicting the effect of mutations on the binding affinity of protein–protein complexes. However, these results were restricted to mutations which did not change the net charge of the side chains due to known difficulties with modeling perturbations involving a change in charge in FEP. Various methods have been proposed to address this problem. Here we apply the co-alchemical water approach to study the efficacy of FEP calculations of charge changing mutations at the protein–protein interface for the antibody–gp120 system investigated previously and three additional complexes. We achieve an overall root mean square error of 1.2 kcal/mol on a set of 106 cases involving a change in net charge selected by a simple suitability filter using side-chain predictions and solvent accessible surface area to be relevant to a biologic optimization project. Reasonable, although less precise, results are also obtained for the 44 more challenging mutations that involve buried residues, which may in some cases require substantial reorganization of the local protein structure, which can extend beyond the scope of a typical FEP simulation. We believe that the proposed prediction protocol will be of sufficient efficiency and accuracy to guide protein engineering projects for which optimization and/or maintenance of a high degree of binding affinity is a key objective.

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Section: Models and Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Relative Binding Affinity Prediction of Charge-Changing Sequence Mutations with FEP in Protein–Protein Interfaces

Clark

Negron

Hauser

et al. 2019

Journal of Molecular Biology

149

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“…Another human PTP, HsSSH1, is activated under oxidative stress due to Cys oxidation of its negative interactor Hs14-3-3ζ (50). The Cys94 of Hs14-3-3ζ was identified as oxidation-sensitive, and its mutation to Ala or Ser causes significant protein destabilization (51). Notably, 3 S-sulfenylated cysteines from the Arabidopsis orthologous proteins, general regulatory factors, align with Cys94 of Hs14-3-3ζ (Fig.…”

Section: S-sulfenylation At Specific Cysteine Sites Affects Protein Fmentioning

confidence: 99%

Mining for protein S-sulfenylation in Arabidopsis uncovers redox-sensitive sites

Huang

Willems

Wei

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

124

115

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Hydrogen peroxide (H2O2) is an important messenger molecule for diverse cellular processes. H2O2 oxidizes proteinaceous cysteinyl thiols to sulfenic acid, also known as S-sulfenylation, thereby affecting the protein conformation and functionality. Although many proteins have been identified as S-sulfenylation targets in plants, site-specific mapping and quantification remain largely unexplored. By means of a peptide-centric chemoproteomics approach, we mapped 1,537 S-sulfenylated sites on more than 1,000 proteins in Arabidopsis thaliana cells. Proteins involved in RNA homeostasis and metabolism were identified as hotspots for S-sulfenylation. Moreover, S-sulfenylation frequently occurred on cysteines located at catalytic sites of enzymes or on cysteines involved in metal binding, hinting at a direct mode of action for redox regulation. Comparison of human and Arabidopsis S-sulfenylation datasets provided 155 conserved S-sulfenylated cysteines, including Cys181 of the Arabidopsis MITOGEN-ACTIVATED PROTEIN KINASE4 (AtMAPK4) that corresponds to Cys161 in the human MAPK1, which has been identified previously as being S-sulfenylated. We show that, by replacing Cys181 of recombinant AtMAPK4 by a redox-insensitive serine residue, the kinase activity decreased, indicating the importance of this noncatalytic cysteine for the kinase mechanism. Altogether, we quantitatively mapped the S-sulfenylated cysteines in Arabidopsis cells under H2O2 stress and thereby generated a comprehensive view on the S-sulfenylation landscape that will facilitate downstream plant redox studies.

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“…that the buried C94 is a determinant for the stability of 14-3-3ζ (Jandova et al 2018). Accordingly, we noted a protein destabilization caused by mutation of C94, observed in the single mutant C94A and in the double (C25A-C94A and C94A-C189A) and triple mutants, which presented a decrease in Tm of 3.2-5.9 o C compared with wild-type (WT)-14-3-3ζ (Table 1).…”

Section: < Fig 2 >mentioning

confidence: 81%

“…14-3-3γ, η and ε were purified as previously reported (Ghorbani et al 2016;Kleppe et al 2014). 14-3-3 cysteine mutants were prepared as previously reported (Jandova et al 2018). TH expression, purification and phosphorylation was performed as reported (Kleppe et al 2014).…”

Section: Protein Expression Purification and Preparation Of Ebselen-treated 14-3-3ζmentioning

confidence: 99%

Cysteine Modification by Ebselen Reduces the Stability and Cellular Levels of 14-3-3 Proteins

et al. 2021

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The 14-3-3 proteins constitute a family of adapter proteins with many binding partners and biological functions, and are considered promising drug targets in cancer and neuropsychiatry. By screening 1280 small-molecule drugs using differential scanning fluorimetry (DSF), we found 15 compounds that decreased the thermal stability of 14-3-3ζ. Among these compounds, ebselen was identified as a covalent, destabilizing ligand of 14-3-3 isoforms ζ, ε, γ and η. Ebselen bonding decreased 14-3-3ζ binding to its partner Ser19-phosphorylated tyrosine hydroxylase. Characterization of site-directed mutants at cysteine residues in 14-3-3ζ (C25, C94, and C189) by DSF and mass spectroscopy revealed covalent modification by ebselen of all cysteines through a selenylsulphide bond. C25 appeared to be the preferential site of ebselen interaction in vitro, whereas modification of C94 was the main determinant for protein destabilization. At therapeutic relevant concentrations ebselen and ebselenoxide caused a decreased 14-3-3 levels in SHSY5Y, accompanied with an increased degradation, most probably by the ubiquitin-dependent proteasome. Moreover, ebselen-treated zebrafish displayed decreased brain 14-3-3 content, a freezing phenotype and reduced mobility, resembling the effects of lithium, consistent with its proposed action as a safer lithium-mimetic drug. Ebselen has recently emerged as a promising drug candidate in several medical areas, such as cancer, neuropsychiatric disorders and infectious diseases, including Covid-19. Its pleiotropic actions are attributed to antioxidant effects and formation of selenosulfides with critical cysteine residues in proteins. Our work indicates that a destabilization of 14-3-3 may affect the protein interaction networks of this protein family, contributing to the therapeutic potential of ebselen.

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Free energy calculations on the stability of the 14-3-3ζ protein

Cited by 9 publications

References 49 publications

Relative Binding Affinity Prediction of Charge-Changing Sequence Mutations with FEP in Protein–Protein Interfaces

Relative Binding Affinity Prediction of Charge-Changing Sequence Mutations with FEP in Protein–Protein Interfaces

Mining for protein S-sulfenylation in Arabidopsis uncovers redox-sensitive sites

Cysteine Modification by Ebselen Reduces the Stability and Cellular Levels of 14-3-3 Proteins

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