Thermodynamic consequences of Tyr to Trp mutations in the cation-π-mediated binding of trimethyllysine by the HP1 chromodomain.
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ABSTRACT:
SUBMITTER: Krone MW
PROVIDER: S-EPMC8152637 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Trimethyllysine (Kme3) reader proteins are targets for inhibition due to their role in mediating gene expression. Although all such reader proteins bind Kme3 in an aromatic cage, the driving force for binding may differ; some readers exhibit evidence for cation-π interactions whereas others do not. We report a general unnatural amino acid mutagenesis approach to quantify the contribution of individual tyrosines to cation binding using the HP1 chromodomain as a model system. We demonstrate that two tyrosines (Y24 and Y48) bind to a Kme3-histone tail peptide via cation-π interactions, but linear free energy trends suggest they do not contribute equally to binding. X-ray structures and computational analysis suggest that the distance and degree of contact between Tyr residues and Kme3 plays an important role in tuning cation-π-mediated Kme3 recognition. Although cation-π interactions have been studied in a number of proteins, this work is the first to utilize direct binding assays, X-ray crystallography, and modeling, to pinpoint factors that influence the magnitude of the individual cation-π interactions.
| S-EPMC6040664 | biostudies-literature
Project description:Histone modifications are implicated in epigenetic inheritance and are of central importance in regulating chromatin structure and gene expression. A prototype example is the trimethylation (Me3) of lysine 9 on histone 3 (H3), which is a readout by an aromatic cage of the chromodomain of heterochromatin-associated protein 1 (HP1) thereby leading to transcriptional repression and heterochromatin formation. Considering that the lysine methylation does not change the charge state of the histone tail and such aromatic-cage mediated recognition of the quaternary ammonium moiety is emerging as the most striking mechanistic commonality for the state-specific recognition of histone lysine methylation, it is of particular interest and importance to understand the physical origin regarding how the aromatic cage distinguishes between the H3K9Me3 mark and its unmethylated counterpart. Here we have simulated relative binding free energies among HP1 chromodomain-H3 tail complexes differing at position 9 of the H3 tail. Our simulated results further confirm the essential role of cation-pi interactions for the binding of a methylated H3 tail by an HP1 chromodomain but indicate that the effect from an electrostatic origin is not dominant in distinguishing between the H3K9Me3 mark and its unmethylated counterpart. Meanwhile, our calculated free energy difference between H3-tert-butyl norleucine 9 and H3-methylnorleucine 9 in their binding to the HP1 clearly reveals the importance of the charge independent interactions for the state-specific readout of histone lysine trimethylation marks.
| S-EPMC2773244 | biostudies-literature
Project description:BACKGROUND:Stem cell differentiation involves major chromatin reorganisation, heterochromatin formation and genomic relocalisation of structural proteins, including heterochromatin protein 1 gamma (HP1?). As the principal reader of the repressive histone marks H3K9me2/3, HP1 plays a key role in numerous processes including heterochromatin formation and maintenance. RESULTS:We find that HP1? is citrullinated in mouse embryonic stem cells (mESCs) and this diminishes when cells differentiate, indicating that it is a dynamically regulated post-translational modification during stem cell differentiation. Peptidylarginine deiminase 4, a known regulator of pluripotency, citrullinates HP1? in vitro. This requires R38 and R39 within the HP1? chromodomain, and the catalytic activity is enhanced by trimethylated H3K9 (H3K9me3) peptides. Mutation of R38 and R39, designed to mimic citrullination, affects HP1? binding to H3K9me3-containing peptides. Using live-cell single-particle tracking, we demonstrate that R38 and R39 are important for HP1? binding to chromatin in vivo. Furthermore, their mutation reduces the residence time of HP1? on chromatin in differentiating mESCs. CONCLUSION:Citrullination is a novel post-translational modification of the structural heterochromatin protein HP1? in mESCs that is dynamically regulated during mESC differentiation. The citrullinated residues lie within the HP1? chromodomain and are important for H3K9me3 binding in vitro and chromatin association in vivo.
| S-EPMC6444592 | biostudies-literature
Project description:Posttranslational modifications of histone proteins regulate gene expression via complex protein-protein and protein-DNA interactions with chromatin. One such modification, the methylation of lysine, has been shown to induce binding to chromodomains in an aromatic cage [Nielsen PR, et al. (2002) Nature 416:103-107]. The binding generally is attributed to the presence of cation-pi interactions between the methylated lysine and the aromatic pocket. However, whether the cationic component of the interaction is necessary for binding in the aromatic cage has not been addressed. In this article, the interaction of trimethyllysine with tryptophan is compared with that of its neutral analog, tert-butylnorleucine (2-amino-7,7-dimethyloctanoic acid), within the context of a beta-hairpin peptide model system. These two side chains have near-identical size, shape, and polarizabilities but differ in their charges. Comparison of the two peptides reveals that the neutral side chain has no preference for interacting with tryptophan, unlike trimethyllysine, which interacts strongly in a defined geometry. In vitro binding studies of the histone 3A peptide containing trimethyllysine or tert-butylnorleucine to HP1 chromodomain indicate that the cationic moiety is critical for binding in the aromatic cage. This difference in binding affinities demonstrates the necessity of the cation-pi interaction to binding with the chromodomain and its role in providing specificity. This article presents an excellent example of synergy between model systems and in vitro studies that allows for the investigation of the key forces that control biomolecular recognition.
| S-EPMC2040873 | biostudies-other
Project description:Crystal structures of Galpha(i) (and closely related family member Galpha(t)) reveal much of what we currently know about G protein structure, including changes which occur in Switch regions. Galpha(t) exhibits a low rate of basal (uncatalyzed) nucleotide exchange and an ordered Switch II region in the GDP-bound state, unlike Galpha(i), which exhibits higher basal exchange and a disordered Switch II region in Galpha(i)GDP structures. Using purified Galpha(i) and Galpha(t), we examined the intrinsic tryptophan fluorescence of these proteins, which reports conformational changes associated with activation and deactivation of Galpha proteins. In addition to the expected enhancement in tryptophan fluorescence intensity, activation of GalphaGDP proteins was accompanied by a modest but notable red shift in tryptophan emission maxima. We identified a cation-pi interaction between tryptophan and arginine residues in the Switch II of Galpha(i) family proteins that mediates the observed red shift in emission maxima. Furthermore, amino-terminal myristoylation of Galpha(i) resulted in a less polar environment for tryptophan residues in the GTPase domain, consistent with an interaction between the myristoylated amino terminus and the GTPase domain of Galpha proteins. These results reveal unique insights into conformational changes which occur upon activation and deactivation of G proteins in solution.
| S-EPMC2788287 | biostudies-literature
Project description:HP1 proteins are central to the assembly and spread of heterochromatin containing histone H3K9 methylation. The chromodomain (CD) of HP1 proteins specifically recognizes the methyl mark on H3 peptides, but the same extent of specificity is not observed within chromatin. The chromoshadow domain of HP1 proteins promotes homodimerization, but this alone cannot explain heterochromatin spread. Using the S. pombe HP1 protein, Swi6, we show that recognition of H3K9-methylated chromatin in vitro relies on an interface between two CDs. This interaction causes Swi6 to tetramerize on a nucleosome, generating two vacant CD sticky ends. On nucleosomal arrays, methyl mark recognition is highly sensitive to internucleosomal distance, suggesting that the CD sticky ends bridge nearby methylated nucleosomes. Strengthening the CD-CD interaction enhances silencing and heterochromatin spread in vivo. Our findings suggest that recognition of methylated nucleosomes and HP1 spread on chromatin are structurally coupled and imply that methylation and nucleosome arrangement synergistically regulate HP1 function.
| S-EPMC3752404 | biostudies-literature
Project description:Combined oxidative phosphorylation (OXPHOS) system deficiencies are a group of mitochondrial disorders that are associated with a range of clinical phenotypes and genetic defects. They occur in approximately 30% of all OXPHOS disorders and around 4% are combined complex I, III and IV deficiencies. In this study we present two mutations in the mitochondrial tRNA(Trp) (MT-TW) and tRNA(Arg) (MT-TR) genes, m.5556G>A and m.10450A>G, respectively, which were detected in two unrelated patients showing combined OXPHOS complex I, III and IV deficiencies and progressive multisystemic diseases. Both mitochondrial tRNA mutations were almost homoplasmic in fibroblasts and muscle tissue of the two patients and not present in controls. Patient fibroblasts showed a general mitochondrial translation defect. The mutations resulted in lowered steady-state levels and altered conformations of the tRNAs. Cybrid cell lines showed similar tRNA defects and impairment of OXPHOS complex assembly as patient fibroblasts. Our results show that these tRNA(Trp) and tRNA(Arg) mutations cause the combined OXPHOS deficiencies in the patients, adding to the still expanding group of pathogenic mitochondrial tRNA mutations.
| S-EPMC2987211 | biostudies-other
Project description:Scope: As a result of population ageing, the number of Alzheimer’s disease (AD) patients has rapidly increased. There are many hypothesises on the pathogenesis of AD, but its detailed molecular mechanism is still unknown, and so no effective preventive or therapeutic measures have been established. Some reports showed a decrease in levels of norepinephrine (NE) has been suspected to be involved in the decline of cognitive function in AD patients and NE concentrations were decreased in postmortem AD patient brains. Tyr-Trp was identified as being the most effective dipeptide in enhancing norepinephrine (NE) synthesis and metabolism. And Tyr-Trp treatment ameliorated the short-term memory dysfunction in AD model mice caused by amyloid beta (Aβ) 25-35. So, the purpose of this study was to investigate the preventive or/and protective effects of Tyr-Trp administration in AD model mice. Methods and results: ddY mice were fed normal diet. After 7 days of feeding, mice were divided into 3 groups (n=10 per group) as follows: the sham group, which received saline administration and distilled water injection; the Aβ group, which received saline administration and Aβ peptides 25–35 injection; and the Aβ+YW group, which received Tyr-Trp administration and Aβ peptides 25–35 injection. Tyr-Trp was orally administered Tyr-Trp (100 mg kg-1 day-1, twice a day), starting 7 days before Aβ peptides injection. The other groups received oral administration of saline (5 mL kg-1). After Aβ peptides injection or sham operation, each groups received every oral administration. DAN microarray showed that Tyrosine hydroxylase, dopa decarboxylase and dopamine receptor D2 mRNA expressions which were downregulated in Aβ group comparing with sham group were upregulated in Aβ+YW group comparing with Aβ group. In addition, quinoid dihydropteridine reductase mRNA expression which wasn’t changed in Aβ group comparing with sham group was upregulated in Aβ+YW group comparing with Aβ group. Conclusions: These results suggest that Tyr-Trp administration might ameliorate the short-term memory dysfunction in AD model mice because of enhancing norepinephrine (NE) synthesis and metabolism through metabolism of both Tyr and Trp.
2020-04-30 | GSE146400 | GEO
Project description:By combining a favorable turn sequence with a turn flanking Trp/Trp interaction and a C-terminal H-bonding interaction between a backbone amide and an i-2 Trp ring, a particularly stable (DeltaG(U) > 7 kJ/mol) truncated hairpin, Ac-WI-(D-Pro-D-Asn)-KWTG-NH(2), results. In this construct and others with a W-(4-residue turn)-W motif in severely truncated hairpins, the C-terminal Trp is the edge residue in a well-defined face-to-edge (FtE) aryl/aryl interaction. Longer hairpins and those with six-residue turns retain the reversed "edge-to-face" (EtF) Trp/Trp geometry first observed for the trpzip peptides. Mutational studies suggest that the W-(4-residue turn)-W interaction provides at least 3 kJ/mol of stabilization in excess of that due to the greater beta-propensity of Trp. The pi-cation, and Trp/Gly-H(N) interactions have been defined. The latter can give rise to >3 ppm upfield shifts for the Gly-H(N) in -WX(n)G- units both in turns (n = 2) and at the C-termini (n = 1) of hairpins. Terminal YTG units result in somewhat smaller shifts (extrapolated to 2 ppm for 100% folding). In peptides with both the EtF and FtE W/W interaction geometries, Trp to Tyr mutations indicate that Trp is the preferred "face" residue in aryl/aryl pairings, presumably because of its greater pi basicity.
| S-EPMC2656586 | biostudies-literature
Project description:Cation-pi interactions in protein structures are identified and evaluated by using an energy-based criterion for selecting significant sidechain pairs. Cation-pi interactions are found to be common among structures in the Protein Data Bank, and it is clearly demonstrated that, when a cationic sidechain (Lys or Arg) is near an aromatic sidechain (Phe, Tyr, or Trp), the geometry is biased toward one that would experience a favorable cation-pi interaction. The sidechain of Arg is more likely than that of Lys to be in a cation-pi interaction. Among the aromatics, a strong bias toward Trp is clear, such that over one-fourth of all tryptophans in the data bank experience an energetically significant cation-pi interaction.
| S-EPMC22230 | biostudies-literature