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1.
J Med Chem ; 63(6): 3172-3187, 2020 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-32125853

RESUMO

Drug resistance has been a major threat in cancer therapies that necessitates the development of new strategies to overcome this problem. We report here a cell-based high-throughput screen of a library containing two-million molecules for the compounds that inhibit the proliferation of non-small-cell lung cancer (NSCLC). Through the process of phenotypic screening, target deconvolution, and structure-activity relationship (SAR) analysis, a compound of furanonaphthoquinone-based small molecule, AS4583, was identified that exhibited potent activity in tyrosine kinase inhibitor (TKI)-sensitive and TKI-resistant NSCLC cells (IC50 = 77 nM) and in xenograft mice. The mechanistic studies revealed that AS4583 inhibited cell-cycle progression and reduced DNA replication by disrupting the formation of the minichromosomal maintenance protein (MCM) complex. Subsequent SAR study of AS4583 gave compound RJ-LC-07-48 which exhibited greater potency in drug-resistant NSCLC cells (IC50 = 17 nM) and in mice with H1975 xenograft tumor.

2.
J Am Chem Soc ; 142(11): 5282-5292, 2020 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-32083852

RESUMO

Human endo-O-sulfatases (Sulf-1 and Sulf-2) are extracellular heparan sulfate proteoglycan (HSPG)-specific 6-O-endosulfatases, which regulate a multitude of cell-signaling events through heparan sulfate (HS)-protein interactions and are associated with the onset of osteoarthritis. These endo-O-sulfatases are transported onto the cell surface to liberate the 6-sulfate groups from the internal d-glucosamine residues in the highly sulfated subdomains of HSPGs. In this study, a variety of HS oligosaccharides with different chain lengths and N- and O-sulfation patterns via chemical synthesis were systematically studied about the substrate specificity of human Sulf-1 employing the fluorogenic substrate 4-methylumbelliferyl sulfate (4-MUS) in a competition assay. The trisaccharide sulfate IdoA2S-GlcNS6S-IdoA2S was found to be the minimal-size substrate for Sulf-1, and substitution of the sulfate group at the 6-O position of the d-glucosamine unit with the sulfonamide motif effectively inhibited the Sulf-1 activity with IC50 = 0.53 µM, Ki = 0.36 µM, and KD = 12 nM.

3.
Sci Rep ; 10(1): 2735, 2020 Feb 17.
Artigo em Inglês | MEDLINE | ID: mdl-32066763

RESUMO

Resistance to cancer therapy is a challenge because of innate tumor heterogeneity and constant tumor evolution. Since the pathway of resistance cannot be predicted, combination therapies may address this progression. We discovered that in addition to IGF1 and IGF2, IGFBP-3 binds bFGF, HGF, neuregulin, and PDGF AB with nanomolar affinity. Because growth factors drive resistance, simultaneous inhibition of multiple growth factor pathways may improve the efficacy of precision therapy. Growth factor sequestration by IGFBP-3-Fc enhances the activity of EGFR inhibitors by decreasing cell survival and inhibiting bFGF, HGF, and IGF1 growth factor rescue and also potentiates the activity of other cancer drugs. Inhibition of tumor growth in vivo with adjuvant IGFBP-3-Fc with erlotinib versus erlotinib after treatment cessation supports that the combination reduces cell survival. Inhibition of multiple growth factor pathways may postpone resistance and extend progression-free survival in many cancer indications.

4.
Genome Biol Evol ; 12(2): 3890-3905, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31971587

RESUMO

Tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis, kills over 1 million people worldwide annually. Development of drug resistance (DR) in the pathogen is a major challenge for TB control. We conducted whole-genome analysis of seven Taiwan M. tuberculosis isolates: One drug susceptible (DS) and five DR Beijing lineage isolates and one DR Euro-American lineage isolate. Developing a new method for DR mutation identification and applying it to the next-generation sequencing (NGS) data from the 6 Beijing lineage isolates, we identified 13 known and 6 candidate DR mutations and provided experimental support for 4 of them. We assembled the genomes of one DS and two DR Beijing lineage isolates and the Euro-American lineage isolate using NGS data. Moreover, using both PacBio and NGS sequencing data, we obtained a high-quality assembly of an extensive DR Beijing lineage isolate. Comparative analysis of these five newly assembled genomes and two published complete genomes revealed a large number of genetic changes, including gene gains and losses, indels and translocations, suggesting rapid evolution of M. tuberculosis. We found the MazEF toxin-antitoxin system in all the seven isolates studied and several interesting mutations in MazEF proteins. Finally, we used the four assembled Beijing lineage genomes to construct a high-quality Beijing lineage reference genome that is DS and contains all the genes in the four genomes. It contains 212 genes not found in the standard reference H37Rv, which is Euro-American. It is therefore a better reference than H37Rv for the Beijing lineage, the predominant lineage in Asia.

5.
J Med Chem ; 63(1): 205-215, 2020 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-31769665

RESUMO

Antiviral drug resistance in influenza infections has been a major threat to public health. To develop a broad-spectrum inhibitor of influenza to combat the problem of drug resistance, we previously identified the highly conserved E339...R416 salt bridge of the nucleoprotein trimer as a target and compound 1 as an inhibitor disrupting the salt bridge with an EC50 = 2.7 µM against influenza A (A/WSN/1933). We have further modified this compound via a structure-based approach and performed antiviral activity screening to identify compounds 29 and 30 with EC50 values of 110 and 120 nM, respectively, and without measurable host cell cytotoxicity. Compared to the clinically used neuraminidase inhibitors, these two compounds showed better activity profiles against drug-resistant influenza A strains, as well as influenza B, and improved survival of influenza-infected mice.

6.
J Vis Exp ; (151)2019 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-31545316

RESUMO

This article presents a general experimental protocol for programmable one-pot oligosaccharide synthesis and demonstrates how to use Auto-CHO software for generating potential synthetic solutions. The programmable one-pot oligosaccharide synthesis approach is designed to empower fast oligosaccharide synthesis of large amounts using thioglycoside building blocks (BBLs) with the appropriate sequential order of relative reactivity values (RRVs). Auto-CHO is a cross-platform software with a graphical user interface that provides possible synthetic solutions for programmable one-pot oligosaccharide synthesis by searching a BBL library (containing about 150 validated and >50,000 virtual BBLs) with accurately predicted RRVs by support vector regression. The algorithm for hierarchical one-pot synthesis has been implemented in Auto-CHO and uses fragments generated by one-pot reactions as new BBLs. In addition, Auto-CHO allows users to give feedback for virtual BBLs to keep valuable ones for further use. One-pot synthesis of stage-specific embryonic antigen 4 (SSEA-4), which is a pluripotent human embryonic stem cell marker, is demonstrated in this work.

7.
Eur J Med Chem ; 181: 111551, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31376567

RESUMO

An 8-oxopurine-6-carboxamide compound (1a) was previously identified as an inhibitor of non-small cell lung cancer (NSCLC). In this study, more than 30 purine-6-carboxamide derivatives with variations at the C2, N7, C8, and N9 positions were synthesized to investigate the structure-activity relationship as a basis for the construction of an advanced pharmacophore model. This model suggests that purine-6-hydroxamate and purine-6-amidoxime analogs could form more hydrogen bonds with a target protein to enhance the inhibitory activities against H1975 cells. Among the series of analogs, hydroxamate 17 and amidoxime 19a exhibited excellent potency against H1975 cells (IC50 < 1.5 µM) and other lung cancer cells with either wild-type or mutated epidermal growth factor receptor (EGFR). Mouse experiments indicated that compounds 17 and 19a were efficient anticancer agents with no appreciable toxicity. The mechanisms of action for the induction of cell apoptosis were determined to involve microtubule fragmentation and p53-mediated signaling pathways.


Assuntos
Amidas/farmacologia , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Ácidos Hidroxâmicos/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Oximas/farmacologia , Purinas/farmacologia , Amidas/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Ácidos Hidroxâmicos/química , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Oximas/química , Purinas/química , Relação Estrutura-Atividade
8.
Eur J Med Chem ; 181: 111584, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31419740

RESUMO

Developing new therapeutic strategies to overcome drug resistance of cancer cells is an ongoing endeavor. From among 2 million chemicals, we identified ethyl 4-oxo-2-phenyl-1,4-dihydroquinoline-6-carboxylate (AS1712) as a low-toxicity inhibitor of lung cancer cell proliferation and xenograft tumor growth. We show that AS1712 is active against broad cancer cell lines and is able to bind in the colchicine-binding pocket of ß-tubulin, thereby inhibiting microtubule assembly and, consequently, inducing mitotic arrest and apoptosis. Our cell-based structure-activity relationship study identified a new lead compound, RJ-LC-15-8, which had a greater anti-proliferative potency for H1975 cells than did AS1712, while maintaining a similar mechanism of action. Notably, AS1712 and RJ-LC-15-8 overcame P-glycoprotein efflux pump and ß-tubulin alterations that lead to acquired resistance against microtubule-targeting drugs of cancer cells. AS1712 and RJ-LC-15-8 may be lead compounds that overcome acquired resistance to microtubule-targeting agents of cancer cells.


Assuntos
Quinolonas/química , Quinolonas/farmacologia , Moduladores de Tubulina/química , Moduladores de Tubulina/farmacologia , Tubulina (Proteína)/metabolismo , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Sítios de Ligação/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colchicina/metabolismo , Resistencia a Medicamentos Antineoplásicos , Humanos , Simulação de Acoplamento Molecular , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Tubulina (Proteína)/química
9.
Biochemistry ; 2019 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-31454239

RESUMO

Carbohydrates make up one of the four major classes of biomolecules, often conjugated with proteins as glycoproteins or with lipids as glycolipids, and participate in many important biochemical functions in living species. However, glycoproteins or glycolipids often exist as mixtures, and as a consequence, it is difficult to isolate individual glycoproteins or glycolipids as pure forms to understand the role carbohydrates play in the glycoconjugate. Currently, the only feasible way to obtain pure glycoconjugates is through synthesis, and of the many methods developed for the synthesis of oligosaccharides, those with automatic and programmable potential are considered to be more effective for addressing the issues of carbohydrate diversity and related functions. In this Perspective, we describe how data science, including algorithm and machine learning, can be used to assist the chemical synthesis of oligosaccharide in a programmable and one-pot manner and how the programmable method can be used to accelerate the construction of diverse oligosaccharides to facilitate our understanding of glycosylation in biology.

10.
ACS Infect Dis ; 5(9): 1493-1504, 2019 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-31283163

RESUMO

One of the major threats to human life nowadays is widespread antibiotic resistance. Antibiotics are used to treat bacterial infections by targeting their essential pathways, such as the biosynthesis of bacterial cell walls. Bacterial transglycosylase, particularly glycosyltransferase family 51 (GT51), is one critical player in the cell wall biosynthesis and has long been known as a promising yet challenging target for antibiotic development. Here, we review the structural studies of this protein and summarize recent progress in developing its specific inhibitors, including synthetic substrate analogs and novel compounds identified from high-throughput screens. A detailed analysis of the protein-ligand interface has also provided us with valuable insights into the future antibiotic development against the bacterial transglycosylase.

11.
J Am Chem Soc ; 141(26): 10309-10314, 2019 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-31244187

RESUMO

Idraparinux is a fully O-sulfated α-methyl glycoside of heparin pentasaccharide motif known to interact with the antithrombin III domain and act as anticoagulant. The current most effective synthesis of Idraparinux is complicated and nonstereoselective, requiring numerous stepwise procedures with low yields. We report here an efficient modular one-pot synthesis of Idraparinux involving the use of a glycosyl phosphate with 6- O- tert-butyl diphenyl silyl group and a d-glucuronic acid-containing disaccharide thioglycoside with 6- O-acetyl group as donor building blocks for the α-directing one-pot glycosylations with an l-iduronic acid-containing disaccharide acceptor building block. The uronic acid was incorporated in a disaccharide module used in the one-pot synthesis to avoid the complicated late-stage installation of these acidic sugars. The one-pot synthesis of Idraparinux demonstrated here is an effective strategy and should be applicable to the modular assembly of other heparan sulfates with regiodefined sulfation pattern for functional study.

12.
J Am Chem Soc ; 141(16): 6484-6488, 2019 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-30969765

RESUMO

Fluorinated glycosides are known to resist the glycosidase-catalyzed glycosidic bond cleavage; however, the synthesis of such glycans, especially 3-fluoro-sialic acid (3F-Neu5Ac) containing sialosides, has been a major challenge. Though the enzymatic synthesis of α-2,3-linked 3F-sialosides was reported, until recently there has not been any effective method available for the synthesis of 3F-sialosides in the α-2,6-linkage. In order to understand the biological effect of such modification, we report here a chemical synthesis of 3Fax-Neu5Ac-α2,6-Gal as a building block for the assembly of 3Fax-Neu5Ac-containing sialosides and a representative homogeneous antibody glycoform. Our results showed that the sialosides are stable under sialidase catalysis and the rituximab glycoform with a sialylated complex-type biantennary glycan terminated with 3Fax-Neu5Ac in the α-2,6-linkage (α2,6-F-SCT) has a similar binding avidity as its parent glycoform. These findings open up new opportunities for the development of therapeutic glycoproteins with improved pharmacokinetic parameters.

13.
Proc Natl Acad Sci U S A ; 116(9): 3518-3523, 2019 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-30808745

RESUMO

The globo-series glycosphingolipids (GSLs) SSEA3, SSEA4, and Globo-H specifically expressed on cancer cells are found to correlate with tumor progression and metastasis, but the functional roles of these GSLs and the key enzyme ß1,3-galactosyltransferase V (ß3GalT5) that converts Gb4 to SSEA3 remain largely unclear. Here we show that the expression of ß3GalT5 significantly correlates with tumor progression and poor survival in patients, and the globo-series GSLs in breast cancer cells form a complex in membrane lipid raft with caveolin-1 (CAV1) and focal adhesion kinase (FAK) which then interact with AKT and receptor-interacting protein kinase (RIP), respectively. Knockdown of ß3GalT5 disrupts the complex and induces apoptosis through dissociation of RIP from the complex to interact with the Fas death domain (FADD) and trigger the Fas-dependent pathway. This finding provides a link between SSEA3/SSEA4/Globo-H and the FAK/CAV1/AKT/RIP complex in tumor progression and apoptosis and suggests a direction for the treatment of breast cancer, as demonstrated by the combined use of antibodies against Globo-H and SSEA4.


Assuntos
Neoplasias da Mama/genética , Galactosiltransferases/genética , Glicoesfingolipídeos/genética , Microdomínios da Membrana/genética , Antígenos Glicosídicos Associados a Tumores/genética , Antígenos Glicosídicos Associados a Tumores/metabolismo , Apoptose/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caveolina 1/genética , Caveolina 1/metabolismo , Progressão da Doença , Proteína de Domínio de Morte Associada a Fas/genética , Feminino , Proteína-Tirosina Quinases de Adesão Focal/genética , Regulação Neoplásica da Expressão Gênica/genética , Glicoesfingolipídeos/metabolismo , Humanos , Substâncias Macromoleculares/química , Substâncias Macromoleculares/metabolismo , Microdomínios da Membrana/metabolismo , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-akt/genética , Saporinas/genética , Transdução de Sinais/genética , Antígenos Embrionários Estágio-Específicos/genética , Antígenos Embrionários Estágio-Específicos/metabolismo
14.
Proc Natl Acad Sci U S A ; 116(10): 4200-4205, 2019 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-30782805

RESUMO

Each year influenza virus infections cause hundreds of thousands of deaths worldwide and a significant level of morbidity with major economic burden. At the present time, vaccination with inactivated virus vaccine produced from embryonated chicken eggs is the most prevalent method to prevent the infections. However, current influenza vaccines are only effective against closely matched circulating strains and must be updated and administered yearly. Therefore, generating a vaccine that can provide broad protection is greatly needed for influenza vaccine development. We have previously shown that vaccination of the major surface glycoprotein hemagglutinin (HA) of influenza virus with a single N-acetylglucosamine at each of the N-glycosylation sites [monoglycosylated HA (HAmg)] can elicit better cross-protection compared with the fully glycosylated HA (HAfg). In the current study, we produced monoglycosylated inactivated split H1N1 virus vaccine from chicken eggs by the N-glycosylation process inhibitor kifunensine and the endoglycosidase Endo H, and intramuscularly immunized mice to examine its efficacy. Compared with vaccination of the traditional influenza vaccine with complex glycosylations from eggs, the monoglycosylated split virus vaccine provided better cross-strain protection against a lethal dose of virus challenge in mice. The enhanced antibody responses induced by the monoglycosylated vaccine immunization include higher neutralization activity, higher hemagglutination inhibition, and more HA stem selectivity, as well as, interestingly, higher antibody-dependent cellular cytotoxicity. This study provides a simple and practical procedure to enhance the cross-strain protection of influenza vaccine by removing the outer part of glycans from the virus surface through modifications of the current egg-based process.

15.
J Am Chem Soc ; 141(9): 3735-3754, 2019 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-30716271

RESUMO

The translation of biological glycosylation in humans to the clinical applications involves systematic studies using homogeneous samples of oligosaccharides and glycoconjugates, which could be accessed by chemical, enzymatic or other biological methods. However, the structural complexity and wide-range variations of glycans and their conjugates represent a major challenge in the synthesis of this class of biomolecules. To help navigate within many methods of oligosaccharide synthesis, this Perspective offers a critical assessment of the most promising synthetic strategies with an eye on the therapeutically relevant targets.

16.
Virology ; 526: 125-137, 2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30388628

RESUMO

The development of a universal influenza vaccine has become a major effort to combat the high mutation rate of influenza. To explore the use of the highly conserved stem region of hemagglutinin (HA) as a universal vaccine, we produced HA-stem-based protein using yeast expression systems. The glycosylation effects on the immunogenicity and protection activities were investigated. The yield of the A/Brisbane/59/2007 HA stem produced from Pichia pastoris reached 100 mg/l. The immunogenicity of HA stem proteins in various glycoforms was further investigated and compared. All glycoforms of the HA stem protein can induce cross-reactive antibody responses, antibody-dependent cellular cytotoxicity (ADCC)-mediated protection as well as T-cell responses, with broad protection in mice. The monoglycosylated form of the A/Brisbane/59/2007 HA stem produced in yeast, together with the glycolipid C34 as the adjuvant, can elicit greater ADCC responses, better neutralizing activities against heterologous strains, and broader protection in mice.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Virus da Influenza A Subtipo H5N1/imunologia , Vacinas contra Influenza/imunologia , Influenza Humana/prevenção & controle , Pichia/metabolismo , Adjuvantes Imunológicos , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Citotoxicidade Celular Dependente de Anticorpos , Técnicas de Cultura Celular por Lotes , Reações Cruzadas , Modelos Animais de Doenças , Feminino , Glicosilação , Glicoproteínas de Hemaglutininação de Vírus da Influenza/biossíntese , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Humanos , Vacinas contra Influenza/biossíntese , Vacinas contra Influenza/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Pichia/genética , Linfócitos T/metabolismo
17.
Nat Commun ; 9(1): 5202, 2018 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-30523255

RESUMO

The programmable one-pot oligosaccharide synthesis method was designed to enable the rapid synthesis of a large number of oligosaccharides, using the software Optimer to search Building BLocks (BBLs) with defined relative reactivity values (RRVs) to be used sequentially in the one-pot reaction. However, there were only about 50 BBLs with measured RRVs in the original library and the method could only synthesize small oligosaccharides due to the RRV ordering requirement. Here, we increase the library to include 154 validated BBLs and more than 50,000 virtual BBLs with predicted RRVs by machine learning. We also develop the software Auto-CHO to accommodate more data handling and support hierarchical one-pot synthesis using fragments as BBLs generated by the one-pot synthesis. This advanced programmable one-pot method provides potential synthetic solutions for complex glycans with four successful examples demonstrated in this work.


Assuntos
Biologia Computacional/métodos , Oligossacarídeos/química , Oligossacarídeos/síntese química , Software , Sequência de Carboidratos , Modelos Químicos , Estrutura Molecular , Polissacarídeos/síntese química , Polissacarídeos/química , Reprodutibilidade dos Testes
18.
Chem Sci ; 9(32): 6685-6691, 2018 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-30310602

RESUMO

Heparin (H) and heparan sulfate (HS) belong to the glycosaminoglycan (GAG) family of oligosaccharides, and their sequences and sulfation patterns are known to regulate the functions of various proteins in biological processes. Among these, the 6-O-sulfation of HS/H contributes most significantly to the structural diversity and binding interactions. However, the synthesis of HS with defined sulfation patterns remains a major challenge. Herein, we report a highly efficient and programmable one-pot method for the synthesis of protected heparin pentasaccharides using thioglycoside building blocks with optimized relative reactivities to allow the selective deprotection and preparation of regiodefined sulfate derivatives.

19.
Chem Commun (Camb) ; 54(56): 7858, 2018 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-29966030

RESUMO

Correction for 'Development of glycosynthases with broad glycan specificity for the efficient glyco-remodeling of antibodies' by Sachin S. Shivatare et al., Chem. Commun., 2018, 54, 6161-6164.

20.
Proc Natl Acad Sci U S A ; 115(28): 7302-7307, 2018 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-29941599

RESUMO

Protein O-glycosylation by attachment of ß-N-acetylglucosamine (GlcNAc) to the Ser or Thr residue is a major posttranslational glycosylation event and is often associated with protein folding, stability, and activity. The methylation of histone H3 at Lys-27 catalyzed by the methyltransferase EZH2 was known to suppress gene expression and cancer development, and we previously reported that the O-GlcNAcylation of EZH2 at S76 stabilized EZH2 and facilitated the formation of H3K27me3 to inhibit tumor suppression. In this study, we employed a fluorescence-based method of sugar labeling combined with mass spectrometry to investigate EZH2 glycosylation and identified five O-GlcNAcylation sites. We also find that mutation of one or more of the O-GlcNAcylation sites S73A, S76A, S84A, and T313A in the N-terminal region decreases the stability of EZH2, but does not affect its association with the PRC2 components SUZ12 and EED. Mutation of the C-terminal O-GlcNAcylation site (S729A) in the catalytic domain of EZH2 abolishes the di- and trimethylation activities, but not the monomethylation of H3K27, nor the integrity of the PRC2/EZH2 core complex. Our results show the effect of individual O-GlcNAcylation sites on the function of EZH2 and suggest an alternative approach to tumor suppression through selective inhibition of EZH2 O-GlcNAcylation.


Assuntos
Acetilglucosamina/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Mutação de Sentido Incorreto , Acetilglucosamina/química , Acetilglucosamina/genética , Substituição de Aminoácidos , Linhagem Celular , Proteína Potenciadora do Homólogo 2 de Zeste/química , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Estabilidade Enzimática , Glicosilação , Humanos , Domínios Proteicos
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