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1.
Chem Commun (Camb) ; 58(87): 12196-12199, 2022 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-36239132

RESUMO

The flexible N-terminal histone tails are a subject of numerous posttranslational modifications, including methylation. We report development of stapled histone peptides bearing trimethyllysine as ligands for epigenetic reader proteins. Stronger or weaker binding affinities have been observed for stapled histone peptides relative to linear histones, indicating that selectivity towards reader proteins can be achieved.


Assuntos
Histonas , Peptídeos , Histonas/metabolismo , Metilação , Peptídeos/metabolismo , Epigênese Genética
2.
Commun Chem ; 5(1)2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36071790

RESUMO

N ε-Methylation of lysine residues in histones plays an essential role in the regulation of eukaryotic transcription. The 'highest' methylation mark, N ε-trimethyllysine, is specifically recognised by N ε-trimethyllysine binding 'reader' domains, and undergoes demethylation, as catalysed by 2-oxoglutarate dependent JmjC oxygenases. We report studies on the recognition of the closest positively charged N ε-trimethyllysine analogue, i.e. its trimethylphosphonium derivative (KPme3), by N ε-trimethyllysine histone binding proteins and Nε-trimethyllysine demethylases. Calorimetric and computational studies with histone binding proteins reveal that H3KP4me3 binds more tightly than the natural H3K4me3 substrate, though the relative differences in binding affinity vary. Studies with JmjC demethylases show that some, but not all, of them can accept the phosphonium analogue of their natural substrates and that the methylation state selectivity can be changed by substitution of nitrogen for phosphorus. The combined results reveal that very subtle changes, e.g. substitution of nitrogen for phosphorus, can substantially affect interactions between ligand and reader domains / demethylases, knowledge that we hope will inspire the development of highly selective small molecules modulating their activity.

3.
Molecules ; 25(8)2020 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-32326252

RESUMO

Gaining a fundamental insight into the biomolecular recognition of posttranslationally modified histones by epigenetic reader proteins is of crucial importance to understanding the regulation of the activity of human genes. Here, we seek to establish whether trimethylthialysine, a simple trimethyllysine analogue generated through cysteine alkylation, is a good trimethyllysine mimic for studies on molecular recognition by reader proteins. Histone peptides bearing trimethylthialysine and trimethyllysine were examined for binding with five human reader proteins employing a combination of thermodynamic analyses, molecular dynamics simulations and quantum chemical analyses. Collectively, our experimental and computational findings reveal that trimethylthialysine and trimethyllysine exhibit very similar binding characteristics for the association with human reader proteins, thereby justifying the use of trimethylthialysine for studies aimed at dissecting the origin of biomolecular recognition in epigenetic processes that play important roles in human health and disease.


Assuntos
Cisteína/análogos & derivados , Histonas/química , Lisina/análogos & derivados , Sítios de Ligação , Cisteína/síntese química , Cisteína/química , Epigênese Genética , Histonas/metabolismo , Humanos , Lisina/síntese química , Lisina/química , Metilação , Modelos Moleculares , Conformação Molecular , Ligação Proteica , Técnicas de Síntese em Fase Sólida , Relação Estrutura-Atividade , Termodinâmica
4.
Commun Chem ; 3(1): 69, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-36703460

RESUMO

The understanding of biomolecular recognition of posttranslationally modified histone proteins is centrally important to the histone code hypothesis. Despite extensive binding and structural studies on the readout of histones, the molecular language by which posttranslational modifications on histone proteins are read remains poorly understood. Here we report physical-organic chemistry studies on the recognition of the positively charged trimethyllysine by the electron-rich aromatic cage containing PHD3 finger of KDM5A. The aromatic character of two tryptophan residues that solely constitute the aromatic cage of KDM5A was fine-tuned by the incorporation of fluorine substituents. Our thermodynamic analyses reveal that the wild-type and fluorinated KDM5A PHD3 fingers associate equally well with trimethyllysine. This work demonstrates that the biomolecular recognition of trimethyllysine by fluorinated aromatic cages is associated with weaker cation-π interactions that are compensated by the energetically more favourable trimethyllysine-mediated release of high-energy water molecules that occupy the aromatic cage.

5.
Bioconjug Chem ; 30(3): 952-958, 2019 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-30794748

RESUMO

Site-specific incorporation of post-translationally modified amino acids into proteins, including histones, has been a subject of great interest for chemical and biochemical communities. Here, we describe a site-specific incorporation of structurally simplest trimethyllysine analogs into position 4 of the intact histone H3 protein. An efficient alkylation of cysteine 4 of the recombinantly expressed histone H3 provides a panel of trimethyllysine analogs that differ in charge, charge density, sterics, and chain length. We demonstrate that H3 histone that bears trimethyllysine analogs can be further assembled into the octameric histone complex that constitutes the nucleosome. Binding studies showed that H3 histone that possesses trimethyllysine analogs is well recognized by a PHD3 reader domain of human JARID1A. This work provides important (bio)chemical tools for fundamental biomolecular studies aimed at unravelling the molecular basis of the higher order nucleosome and chromatin assemblies.


Assuntos
Cisteína/química , Histonas/química , Lisina/análogos & derivados , Alquilação , Animais , Eletroforese em Gel de Poliacrilamida , Histonas/metabolismo , Humanos , Lisina/química , Processamento de Proteína Pós-Traducional , Proteína 2 de Ligação ao Retinoblastoma/metabolismo , Espectrometria de Massas por Ionização por Electrospray , Xenopus laevis
6.
Mol Plant ; 11(8): 1038-1052, 2018 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-29793052

RESUMO

Heterochromatin Protein 1 (HP1) is a major regulator of chromatin structure and function. In animals, the network of proteins interacting with HP1 is mainly associated with constitutive heterochromatin marked by H3K9me3. HP1 physically interacts with the putative ortholog of the SNF2 chromatin remodeler ATRX, which controls deposition of histone variant H3.3 in mammals. In this study, we show that the Arabidopsis thaliana ortholog of ATRX participates in H3.3 deposition and possesses specific conserved domains in plants. We found that plant Like HP1 (LHP1) protein interacts with ATRX through domains that evolved specifically in land plant ancestors. Loss of ATRX function in Arabidopsis affects the expression of a limited subset of genes controlled by PRC2 (POLYCOMB REPRESSIVE COMPLEX 2), including the flowering time regulator FLC. The function of ATRX in regulation of flowering time requires novel LHP1-interacting domain and ATPase activity of the ATRX SNF2 helicase domain. Taken together, these results suggest that distinct evolutionary pathways led to the interaction between ATRX and HP1 in mammals and its counterpart LHP1 in plants, resulting in distinct modes of transcriptional regulation.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Repressoras/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Homólogo 5 da Proteína Cromobox , Proteínas Cromossômicas não Histona/genética , Histonas/metabolismo , Complexo Repressor Polycomb 2 , Proteínas Repressoras/genética
7.
Chem Commun (Camb) ; 54(19): 2409-2412, 2018 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-29457186

RESUMO

Histone Nε-lysine methylation is a widespread posttranslational modification that is specifically recognised by a diverse class of Nε-methyllysine binding reader proteins. Combined thermodynamic data, molecular dynamics simulations, and quantum chemical studies reveal that reader proteins efficiently bind trimethylornithine and trimethylhomolysine, the simplest Nε-trimethyllysine analogues that differ in the length of the side chain.


Assuntos
Proteínas de Transporte/química , Epigênese Genética , Histonas/química , Lisina/análogos & derivados , Fragmentos de Peptídeos/química , Proteínas de Transporte/genética , Histonas/genética , Humanos , Lisina/química , Lisina/genética , Lisina/metabolismo , Simulação de Dinâmica Molecular , Estrutura Molecular , Ornitina/análogos & derivados , Fragmentos de Peptídeos/genética , Ligação Proteica , Teoria Quântica , Termodinâmica
8.
Chem Commun (Camb) ; 53(99): 13264-13267, 2017 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-29186216

RESUMO

Histone lysine methylation is regulated by Nε-methyltransferases, demethylases, and Nε-methyl lysine binding proteins. Thermodynamic, catalytic and computational studies were carried out to investigate the interaction of three epigenetic protein classes with synthetic histone substrates containing l- and d-lysine residues. The results reveal that out of the three classes, Nε-methyl lysine binding proteins are superior in accepting lysines with the d-configuration.


Assuntos
Epigênese Genética/genética , Histona Desmetilases/metabolismo , Lisina/química , Metiltransferases/metabolismo , Biocatálise , Histona Desmetilases/genética , Lisina/metabolismo , Metilação , Metiltransferases/genética , Modelos Moleculares , Conformação Molecular , Estereoisomerismo , Termodinâmica
9.
Chem Commun (Camb) ; 52(87): 12849-12852, 2016 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-27730239

RESUMO

Trimethyllysine hydroxylase (TMLH) is a non-haem Fe(ii) and 2-oxoglutarate dependent oxygenase that catalyses the C-3 hydroxylation of an unactivated C-H bond in l-trimethyllysine in the first step of carnitine biosynthesis. The examination of trimethyllysine analogues as substrates for human TMLH reveals that the enzyme does hydroxylate substrates other than natural l-trimethyllysine.

10.
Chem Soc Rev ; 45(1): 24-39, 2016 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-26497225

RESUMO

Supramolecular protein assemblies are an emerging area within the chemical sciences, which combine the topological structures of the field of supramolecular chemistry and the state-of-the-art chemical biology approaches to unravel the formation and function of protein assemblies. Recent chemical and biological studies on natural multimeric protein structures, including fibers, rings, tubes, catenanes, knots, and cages, have shown that the quaternary structures of proteins are a prerequisite for their highly specific biological functions. In this review, we illustrate that a striking structural diversity of protein assemblies is present in nature. Furthermore, we describe structure-function relationship studies for selected classes of protein architectures, and we highlight the techniques that enable the characterisation of supramolecular protein structures.


Assuntos
Produtos Biológicos/síntese química , Complexos Multiproteicos/síntese química , Animais , Produtos Biológicos/química , Humanos , Modelos Moleculares , Complexos Multiproteicos/química
11.
Nat Commun ; 6: 8911, 2015 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-26578293

RESUMO

A large number of structurally diverse epigenetic reader proteins specifically recognize methylated lysine residues on histone proteins. Here we describe comparative thermodynamic, structural and computational studies on recognition of the positively charged natural trimethyllysine and its neutral analogues by reader proteins. This work provides experimental and theoretical evidence that reader proteins predominantly recognize trimethyllysine via a combination of favourable cation-π interactions and the release of the high-energy water molecules that occupy the aromatic cage of reader proteins on the association with the trimethyllysine side chain. These results have implications in rational drug design by specifically targeting the aromatic cage of readers of trimethyllysine.


Assuntos
Acetiltransferases/química , Antígenos Nucleares/química , Histonas/metabolismo , Histona Desmetilases com o Domínio Jumonji/química , Lisina/análogos & derivados , Proteínas do Tecido Nervoso/química , Proteína 2 de Ligação ao Retinoblastoma/química , Fator de Transcrição TFIID/química , Fatores de Transcrição/química , Aminoácidos Aromáticos/química , Calorimetria , Cristalografia por Raios X , Epigênese Genética , Código das Histonas , Humanos , Lisina/química , Metilação , Modelos Moleculares , Simulação de Dinâmica Molecular , Compostos de Amônio Quaternário , Fatores Associados à Proteína de Ligação a TATA
12.
PLoS One ; 10(9): e0139205, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26421618

RESUMO

Several reader domain proteins that specifically recognize methyllysine-containing histones contain the negatively-charged aspartate or glutamate residues as part of the aromatic cage. Herein, we report thermodynamic analyses for the recognition of histone H3K4me3 and H3K4me2 by the tandem tudor domain of Sgf29 and its recognition site variants. Small uncharged and large aromatic substitutions on the Asp266 site resulted in a significant decrease in binding affinities for both H3K4me3 and H3K4me2, demonstrating the role of the negative charge of Asp266 in the readout process by Sgf29. This study emphasizes the essential contribution of electrostatic interactions to the overall binding affinity, and reveals that the underlying mechanisms for the recognition of Kme2/3 depend on the composition and arrangement of the aromatic cage.


Assuntos
Acetiltransferases/química , Acetiltransferases/metabolismo , Histonas/química , Histonas/metabolismo , Lisina/metabolismo , Eletricidade Estática , Ácido Aspártico/metabolismo , Metilação , Modelos Moleculares , Ligação Proteica , Estrutura Terciária de Proteína , Termodinâmica
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