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1.
J Am Chem Soc ; 2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38848460

RESUMO

Cancer immune therapies, particularly programmed cell death protein 1 (PD-1) blockade immunotherapy, falter in aged individuals due to compromised T-cell immunity. Spermidine, a biogenic polyamine that declines along with aging, shows promise in restoring antitumor immunity by enhancing mitochondrial fatty acid oxidation (FAO). Herein, we report a spermidine-based chemoproteomic probe (probe 2) that enables profiling of spermidine-binding proteins and screening for small-molecule enhancers of mitochondrial FAO. Chemoproteomic profiling by the probe revealed 140 proteins engaged in cellular interaction with spermidine, with a significant majority being mitochondrial proteins. Hydroxyl coenzyme A (CoA) dehydrogenase subunits α (HADHA) and other lipid metabolism-linked proteins are among the mitochondrial proteins that have attracted considerable interest. Screening spermidine analogs with the probe led to the discovery of compound 13, which interacts with these lipid metabolism-linked proteins and activates HADHA. This simple and biostable synthetic compound we named "spermimic" mirrors spermidine's ability to enhance mitochondrial bioenergetics and displays similar effectiveness in augmenting PD-1 blockade therapy in mice. This study lays the foundation for developing small-molecule activators of antitumor immunity, offering potential in combination cancer immunotherapy.

2.
Nucleic Acids Res ; 50(14): 8143-8153, 2022 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-35801908

RESUMO

Tandem repeats of guanine-rich sequences in RNA often form thermodynamically stable four-stranded RNA structures. Such RNA G-quadruplexes have long been considered to be linked to essential biological processes, yet their physiological significance in cells remains unclear. Here, we report a approach that permits the detection of RNA G-quadruplex structures that modulate protein translation in mammalian cells. The approach combines antibody arrays and RGB-1, a small molecule that selectively stabilizes RNA G-quadruplex structures. Analysis of the protein and mRNA products of 84 cancer-related human genes identified Nectin-4 and CapG as G-quadruplex-controlled genes whose mRNAs harbor non-canonical G-quadruplex structures on their 5'UTR region. Further investigations revealed that the RNA G-quadruplex of CapG exhibits a structural polymorphism, suggesting a possible mechanism that ensures the translation repression in a KCl concentration range of 25-100 mM. The approach described in the present study sets the stage for further discoveries of RNA G-quadruplexes.


Assuntos
Quadruplex G , Regiões 5' não Traduzidas , Animais , Guanina/química , Humanos , Mamíferos/genética , Biossíntese de Proteínas , RNA Mensageiro/metabolismo
3.
Nucleic Acids Res ; 49(22): e132, 2021 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-34581825

RESUMO

Imaging the dynamics of proteins in living cells is a powerful means for understanding cellular functions at a deeper level. Here, we report a versatile method for spatiotemporal imaging of specific endogenous proteins in living mammalian cells. The method employs a bifunctional aptamer capable of selective protein recognition and fluorescent probe-binding, which is induced only when the aptamer specifically binds to its target protein. An aptamer for ß-actin protein preferentially recognizes its monomer forms over filamentous forms, resulting in selective G-actin staining in both fixed and living cells. Through actin-drug treatment, the method permitted direct monitoring of the intracellular concentration change of endogenous G-actin. This protein-labeling method, which is highly selective and non-covalent, provides rich insights into the study of spatiotemporal protein dynamics in living cells.


Assuntos
Aptâmeros de Nucleotídeos , Imagem Óptica/métodos , Proteínas/análise , Actinas/análise , Aptâmeros de Nucleotídeos/química , Corantes Fluorescentes , Células HeLa , Humanos , Imagem Molecular/métodos , RNA/química , Imagem com Lapso de Tempo
4.
J Am Chem Soc ; 144(44): 20171-20176, 2022 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-36306265

RESUMO

Visible light, particularly in the blue region of the spectrum, can cause cell dysfunction through the generation of singlet oxygen, contributing to cellular aging and age-related pathologies. Although photooxidation of nucleic acids, lipids, and amino acids has been extensively studied, the magnitude and span of blue-light-induced protein damages within proteome remain largely unknown. Herein we present a chemoproteomic approach to mapping blue-light-damaged proteins in live mammalian cells by exploiting a nucleophilic alkyne chemical probe. A gene ontology enrichment analysis revealed that cell surface proteins are more readily oxidized than other susceptible sets of proteins, including mitochondrial proteins. In particular, the integrin family of cell surface receptors (ITGs) was highly ranked in the mammalian cells tested, including human corneal endothelial cells. The blue-light-oxidized ITGB1 protein was functionally inactive in promoting cell adhesion and proliferation, suggesting that the photodamage of integrins contributes to the blue-light-induced cell dysfunction. Further application of our method to various cells and tissues should lead to a comprehensive analysis of light-sensitive proteins.


Assuntos
Células Endoteliais , Oxigênio Singlete , Animais , Humanos , Oxirredução , Luz , Mamíferos
5.
Cancer Sci ; 113(10): 3608-3617, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-36068652

RESUMO

To overcome the increasing burden on pathologists in diagnosing gastric biopsies, we developed an artificial intelligence-based system for the pathological diagnosis of gastric biopsies (AI-G), which is expected to work well in daily clinical practice in multiple institutes. The multistage semantic segmentation for pathology (MSP) method utilizes the distribution of feature values extracted from patches of whole-slide images (WSI) like pathologists' "low-power view" information of microscopy. The training dataset included WSIs of 4511 gastric biopsy tissues from 984 patients. In tissue-level validation, MSP AI-G showed better accuracy (91.0%) than that of conventional patch-based AI-G (PB AI-G) (89.8%). Importantly, MSP AI-G unanimously achieved higher accuracy rates (0.946 ± 0.023) than PB AI-G (0.861 ± 0.078) in tissue-level analysis, when applied to the cohorts of 10 different institutes (3450 samples of 1772 patients in all institutes, 198-555 samples of 143-206 patients in each institute). MSP AI-G had high diagnostic accuracy and robustness in multi-institutions. When pathologists selectively review specimens in which pathologist's diagnosis and AI prediction are discordant, the requirement of a secondary review process is significantly less compared with reviewing all specimens by another pathologist.


Assuntos
Inteligência Artificial , Estômago , Biópsia , Humanos
6.
Mol Phylogenet Evol ; 175: 107460, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35358691

RESUMO

The Lingulidae are often considered living fossils, because they have shown little morphological change since the Paleozoic. Limited morphological variation has also made the taxonomic study of living lingulids challenging. We investigated species diversity and phylogenetic relationships of extant lingulids and show that they are substantially more diverse than realized, demonstrating that morphological stasis was commonly accompanied by speciation. Species delimitation based on cytochrome c oxidase subunit I (COI) gene sequences from 194 specimens sampled from East Asia, Australia, Oceania, and the Americas suggested 14-22 species in the lingulids (9-17 species in Lingula and 4-5 species in Glottidia), in contrast to the 11-12 species currently recognized globally in the family. Four-gene phylogenetic analyses supported the sister relationship between Lingula and Glottidia. Within Lingula, L. adamsi, which possesses large, brownish shells, was recovered as sister to all remaining Lingula species, which have more or less greenish shells. Within the greenish Lingula clade, the 'L. anatina' complex was sister to the clade that includes the 'L. reevei' complex. The 'L. anatina' complex was further separated into two major clades with partly separate ranges centered on (i) temperate East Asia, and (ii) the tropical west-central Pacific. Within Glottidia, Pacific species were nested within Atlantic species. Time-calibrated phylogenetic analyses suggested that Lingula likely originated in the early Cretaceous contrary to a previously proposed hypothesis advocating a Cenozoic origin. The separation of Lingula and Glottidia appears to date from the Mesozoic, not from the Carboniferous, contrary to a previous hypothesis. Overall, our results uncovered substantial cryptic diversity in lingulids, which will form the basis for conservation and further taxonomic revision.


Assuntos
Fósseis , Hidrozoários , Animais , Ásia Oriental , Invertebrados/genética , Filogenia
7.
J Am Chem Soc ; 142(3): 1142-1146, 2020 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-31899620

RESUMO

We accidentally found that YM-53601, a known small-molecule inhibitor of squalene synthase (SQS), selectively depletes SQS from mammalian cells upon UV irradiation. Further analyses indicated that the photodepletion of SQS requires its short peptide segment located at the COOH terminus. Remarkably, when the 27 amino acid peptide was fused to green fluorescent protein or unrelated proteins at either the NH2 or COOH terminus, such fusion proteins were selectively depleted when the cells were treated with both YM-53601 and UV exposure. Product analysis and electron spin resonance experiments suggested that the UV irradiation promotes homolytic C-O bond cleavage of the aryl ether group in YM-53601. It is likely that the radical species generated from UV-activated YM-53601 abstract hydrogen atoms from the SQS peptide, leading to the photolysis of the entire protein. The pair of the SQS peptide and YM-53601 discovered in the present study paves the way for the design of a new small-molecule-controlled optogenetic tool.


Assuntos
Farnesil-Difosfato Farnesiltransferase/antagonistas & inibidores , Peptídeos/farmacologia , Fotólise , Quinuclidinas/farmacologia , Células HEK293 , Humanos
8.
Angew Chem Int Ed Engl ; 56(7): 1765-1770, 2017 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-28067441

RESUMO

A major hurdle in stem cell therapy is the tumorigenic risk of residual undifferentiated stem cells. This report describes the design and evaluation of synthetic hybrid molecules that efficiently reduce the number of human induced pluripotent stem cells (hiPSCs) in cell mixtures. The design takes advantage of Kyoto probe 1 (KP-1), a fluorescent chemical probe for hiPSCs, and clinically used anticancer drugs. Among the KP-1-drug conjugates we synthesized, we found an exceptionally selective, chemically tractable molecule that induced the death of hiPSCs. Mechanistic analysis suggested that the high selectivity originates from the synergistic combination of transporter-mediated efflux and the cytotoxicity mode of action. The present study offers a chemical and mechanistic rationale for designing selective, safe, and simple reagents for the preparation of non-tumorigenic clinical samples.


Assuntos
Antineoplásicos/química , Separação Celular/métodos , Corantes Fluorescentes/química , Células-Tronco Pluripotentes Induzidas/citologia , Rodaminas/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular , Corantes Fluorescentes/farmacologia , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Rodaminas/farmacologia
9.
J Am Chem Soc ; 138(29): 9037-40, 2016 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-27410677

RESUMO

The G-quadruplexes form highly stable nucleic acid structures, which are implicated in various biological processes in both DNA and RNA. Although DNA G-quadruplexes have been studied in great detail, biological roles of RNA G-quadruplexes have received less attention. Here, a screening of a chemical library permitted identification of a small-molecule tool that binds selectively to RNA G-quadruplex structures. The polyaromatic molecule, RGB-1, stabilizes RNA G-quadruplex, but not DNA versions or other RNA structures. RGB-1 intensified the G-quadruplex-mediated inhibition of RNA translation in mammalian cells, decreased expression of the NRAS proto-oncogene in breast cancer cells, and permitted identification of a novel sequence that forms G-quadruplex in NRAS mRNA. RGB-1 may serve as a unique tool for understanding cellular roles of RNA G-quadruplex structures.


Assuntos
Quadruplex G , Biossíntese de Proteínas/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Avaliação Pré-Clínica de Medicamentos , RNA Mensageiro/química , RNA Mensageiro/genética
10.
BMC Psychiatry ; 16: 66, 2016 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-26980460

RESUMO

BACKGROUND: Bitopertin, a glycine reuptake inhibitor, was investigated as a novel treatment for schizophrenia. We report all the results of a double-blind randomized study assessing safety and efficacy following 52-week adjunctive treatment with bitopertin in Japanese patients with schizophrenia. METHODS: This study enrolled Japanese outpatients with schizophrenia who met criteria for either "negative symptoms", i.e., patients with persistent, predominant negative symptoms of schizophrenia even after long-term treatment with antipsychotics or "sub-optimally controlled symptoms", i.e., patients with insufficiently improved symptoms of schizophrenia even after long-term treatment with antipsychotics, respectively. One hundred sixty-one patients were randomly assigned to receive 52-week treatments with bitopertin doses of 5, 10, or 20 mg/day at ratio of 1:5:5, where existing antipsychotics were concomitantly administered. Efficacy endpoints included Positive and Negative Syndrome Scale (PANSS), Clinical Global Impression (CGI), and Personal and Social Performance (PSP). The purpose of the present study is primarily to evaluate the safety, and secondarily to investigate the clinical efficacy of bitopertin. RESULTS: One hundred fourteen patients (71 %) completed 52-week treatment with bitopertin. Most of the adverse events were mild or moderate in their severity. The patients in the 20-mg group experienced more adverse events than the patients in the other two groups. Common dose-dependent adverse events were somnolence and insomnia associated with worsening schizophrenia. The blood hemoglobin levels gradually decreased from baseline in a dose-dependent manner, but there were no patients with the decrease below 10 g/dL that would have led to their discontinuation. All the efficacy endpoints gradually improved in all the treatment groups for both of the two symptoms, while there were no clear differences among the three dose groups. CONCLUSIONS: Altogether, bitopertin was found to be generally safe and well-tolerated for the treatment of patients with schizophrenia. All three bitopertin treated groups showed improvements in all the efficacy endpoints for both of the two symptoms, i.e., "negative symptoms" and "sub-optimally controlled symptoms", throughout the duration of the study. TRIAL REGISTRATION: Japan Pharmaceutical Information Center, number JapicCTI-111627 (registered on September 20, 2011).


Assuntos
Piperazinas/uso terapêutico , Esquizofrenia/tratamento farmacológico , Sulfonas/uso terapêutico , Adulto , Antipsicóticos/uso terapêutico , Método Duplo-Cego , Feminino , Humanos , Japão , Masculino , Pessoa de Meia-Idade , Resultado do Tratamento
11.
J Am Chem Soc ; 137(50): 15624-7, 2015 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-26632868

RESUMO

Synthetic agents that disrupt intracellular protein-protein interactions (PPIs) are highly desirable for elucidating signaling networks and developing new therapeutics. However, designing cell-penetrating large molecules equipped with the many functional groups necessary for binding to large interfaces remains challenging. Here, we describe a rational strategy for the intracellular oxime ligation-mediated generation of an amphipathic bivalent inhibitor composed of a peptide and diterpene natural product, fusicoccin, which binds 14-3-3 protein with submicromolar affinity. Our results demonstrate that co-treatment of cells with small module molecules, the aldehyde-containing fusicoccin 1 and the aminooxy-containing peptide 2, generates the corresponding conjugate 3 in cells, resulting in significant cytotoxicity. In contrast, chemically synthesized 3 is not cytotoxic, likely due to its inability to penetrate cells. Compound 3, but not 1 or 2, disrupts endogenous 14-3-3/cRaf interactions, suggesting that cell death is caused by inhibition of 14-3-3 activity. These results suggest that intracellular generation of large-sized molecules may serve as a new approach for modulating PPIs.


Assuntos
Proteínas 14-3-3/química , Diterpenos/química , Peptídeos/química , Células HEK293 , Humanos , Simulação de Acoplamento Molecular , Ligação Proteica
12.
J Acoust Soc Am ; 138(2): 769-81, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26328693

RESUMO

Opera houses represent a large group of performance spaces characterized by great complexity and, at the same time, versatility with respect to different usage (from opera to symphonic music and ballet). This kind of building originated in Italy during the 17th century and later spread across the country and then Europe and the rest of the world, slowly evolving into modern theatre shapes. As a consequence of the changes undergone by the interior space, the original acoustic features, which likely influenced many composers, experienced important variations. Thanks to acoustic measurement campaigns inside Italian Historical Opera Houses, promoted by National and Regional Projects, the distinctive features of these spaces were investigated in comparison to modern spaces. In this work, the newly acquired data are merged with data in the literature in order to present and discuss some of the distinctive acoustic features of historical spaces as regards their original function. Moreover, specific issues such as listening in stalls and boxes and the criteria governing the preference judgment of listeners are considered. The concept and the crucial role of the balance between stage and pit sources are also discussed by means of previous literature studies.

13.
Angew Chem Int Ed Engl ; 54(6): 1855-8, 2015 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-25537455

RESUMO

Determination of subcellular localization and dynamics of mRNA is increasingly important to understanding gene expression. A new convenient and versatile method is reported that permits spatiotemporal imaging of specific non-engineered RNAs in living cells. The method uses transfection of a plasmid encoding a gene-specific RNA aptamer, combined with a cell-permeable synthetic small molecule, the fluorescence of which is restored only when the RNA aptamer hybridizes with its cognitive mRNA. The method was validated by live-cell imaging of the endogenous mRNA of ß-actin. Application of the technology to mRNAs of a total of 84 human cytoskeletal genes allowed us to observe cellular dynamics of several endogenous mRNAs including arfaptin-2, cortactin, and cytoplasmic FMR1-interacting protein 2. The RNA-imaging technology and its further optimization might permit live-cell imaging of any RNA molecules.


Assuntos
RNA Mensageiro/genética , Sequência de Bases , Dados de Sequência Molecular
14.
Angew Chem Int Ed Engl ; 53(42): 11208-13, 2014 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-25196666

RESUMO

One of the major problems encountered in cell transplantation is the low level of survival of transplanted cells due to detachment-induced apoptosis, called anoikis. The present study reports on the chemical synthesis and biological evaluation of water-soluble molecules that protect suspended cells from anoikis. The synthetic molecules bind to and induce clusters of integrins and heparan-sulfate-bound syndecans, two classes of receptors that are important for extracellular matrix-mediated cell survival. Molecular biological analysis indicates that such molecules prolong the survival of suspended NIH3T3 cells, at least in part, by promoting clustering of syndecan-4 and integrin ß1 on the cell surface, leading to the activation of small GTPase Rac-1 and Akt. In vivo experiments using animal disease models demonstrated the ability of the molecules to improve cell engraftment. The cluster-inducing molecules may provide a starting point for the design of new synthetic tools for cell-based therapy.


Assuntos
Anoikis/efeitos dos fármacos , Transplante de Células , Peptídeos/química , Peptídeos/farmacologia , Substâncias Protetoras/química , Substâncias Protetoras/farmacologia , Animais , Adesão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transplante de Células/métodos , Células Cultivadas , Integrina beta1/metabolismo , Camundongos , Células NIH 3T3 , Coelhos , Sindecana-4/metabolismo
15.
J Am Chem Soc ; 135(30): 11032-9, 2013 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-23822587

RESUMO

Adhesamine is an organic small molecule that promotes adhesion and growth of cultured human cells by binding selectively to heparan sulfate on the cell surface. The present study combined chemical, physicochemical, and cell biological experiments, using adhesamine and its analogues, to examine the mechanism by which this dumbbell-shaped, non-peptidic molecule induces physiologically relevant cell adhesion. The results suggest that multiple adhesamine molecules cooperatively bind to heparan sulfate and induce its assembly, promoting clustering of heparan sulfate-bound syndecan-4 on the cell surface. A pilot study showed that adhesamine improved the viability and attachment of transplanted cells in mice. Further studies of adhesamine and other small molecules could lead to the design of assembly-inducing molecules for use in cell biology and cell therapy.


Assuntos
Heparitina Sulfato/metabolismo , Piperazinas/química , Piperazinas/farmacologia , Animais , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Dimerização , Desenho de Fármacos , Humanos , Masculino , Camundongos , Modelos Moleculares , Piperazinas/metabolismo , Multimerização Proteica/efeitos dos fármacos , Estrutura Quaternária de Proteína , Relação Estrutura-Atividade , Sindecanas/química
16.
J Biol Chem ; 286(5): 4003-10, 2011 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-21123173

RESUMO

Signal transducer and activator of transcription 6 (STAT6), which plays a critical role in immune responses, is activated by interleukin-4 (IL-4). Activity of STAT family members is regulated primarily by tyrosine phosphorylations and possibly also by serine phosphorylations. Here, we report a previously undescribed serine phosphorylation of STAT6, which is activated by cell stress or by the pro-inflammatory cytokine, interleukin-1ß (IL-1ß). Our analyses suggest that Ser-707 is phosphorylated by c-Jun N-terminal kinase (JNK). Phosphorylation decreases the DNA binding ability of IL-4-stimulated STAT6, thereby inhibiting the transcription of STAT6-responsive genes. Inactivation of STAT6 by JNK-dependent Ser-707 phosphorylation may be one mechanism of controlling the balance between IL-1ß and IL-4 signals.


Assuntos
Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fosforilação/fisiologia , Fator de Transcrição STAT6/metabolismo , Serina/metabolismo , DNA/metabolismo , Ativação Enzimática , Humanos , Interleucina-1beta , Interleucina-4 , Fator de Transcrição STAT6/genética , Estresse Fisiológico , Transcrição Gênica
17.
J Acoust Soc Am ; 131(2): 1206-20, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22352495

RESUMO

In 1965, the Catholic Church liturgy changed to allow priests to face the congregation. Whereas Church tradition, teaching, and participation have been much discussed with respect to priest orientation at Mass, the acoustical changes in this regard have not yet been examined scientifically. To discuss acoustic desired within churches, it is necessary to know the acoustical characteristics appropriate for each phase of the liturgy. In this study, acoustic measurements were taken at various source locations and directions using both old and new liturgies performed in Japanese churches. A directional loudspeaker was used as the source to provide vocal and organ acoustic fields, and impulse responses were measured. Various acoustical parameters such as reverberation time and early decay time were analyzed. The speech transmission index was higher for the new Catholic liturgy, suggesting that the change in liturgy has improved speech intelligibility. Moreover, the interaural cross-correlation coefficient and early lateral energy fraction were higher and lower, respectively, suggesting that the change in liturgy has made the apparent source width smaller.


Assuntos
Acústica , Localização de Som/fisiologia , Catolicismo , Humanos , Decoração de Interiores e Mobiliário , Japão , Espectrografia do Som , Percepção Espacial/fisiologia , Inteligibilidade da Fala/fisiologia
18.
ACS Chem Biol ; 17(2): 340-347, 2022 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-35076225

RESUMO

Covalent inhibitors of enzymes are increasingly appreciated as pharmaceutical seeds, yet discovering non-cysteine-targeting inhibitors remains challenging. Herein, we report an intriguing experience during our activity-based proteomic screening of 1601 reactive small molecules, in which we monitored the ability of library molecules to compete with a cysteine-reactive iodoacetamide probe. One epoxide molecule, F8, exhibited unexpected enhancement of the probe reactivity for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a rate-limiting glycolysis enzyme. In-depth mechanistic analysis suggests that F8 forms a covalent adduct with an aspartic acid in the active site to displace NAD+, a cofactor of the enzyme, with concomitant enhancement of the probe reaction with the catalytic cysteine. The mechanistic underpinning permitted the identification of an optimized aspartate-reactive GAPDH inhibitor. Our findings exemplify that activity-based proteomic screening with a cysteine-reactive probe can be used for discovering covalent inhibitors that react with non-cysteine residues.


Assuntos
Cisteína , Proteômica , Catálise , Domínio Catalítico , Cisteína/química , Gliceraldeído-3-Fosfato Desidrogenases/química , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo
19.
ACS Chem Biol ; 17(3): 567-575, 2022 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-35188733

RESUMO

The present study reports a surprising protein-condensing effect of glucose, prompted by our accidental observation during chemical library screening under a high-glucose condition. We noticed "glucosing-out" of certain compounds, in which physiological concentrations of glucose induced compound aggregation. Adapting the "glucosing-out" concept to proteins, our proteomic analysis identified three cellular proteins (calmodulin, rho guanine nucleotide exchange factor 40, and polyubiquitin-C) that displayed robust glucose-dependent precipitation. One of these proteins, calmodulin, formed glucose-dependent condensates that control cellular glycogenolysis in hepatic cells. Our findings suggest that glucose is a heretofore underappreciated driver of protein phase separation that may have profound effects on cellular homeostasis.


Assuntos
Glucose , Glicogenólise , Calmodulina/metabolismo , Glucose/metabolismo , Homeostase , Proteômica
20.
Chem Sci ; 13(19): 5760-5766, 2022 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-35694339

RESUMO

Phase-separated membraneless organelles or biomolecular condensates play diverse functions in cells, however recapturing their characteristics using small organic molecules has been a challenge. In the present study, cell-lysate-based screening of 843 self-assembling small molecules led to the discovery of a simple organic molecule, named huezole, that forms liquid droplets to selectively sequester tubulin. Remarkably, this small molecule enters cultured human cells and prevents cell mitosis by forming tubulin-concentrating condensates in cells. The present study demonstrates the feasibility of producing a synthetic condensate out of non-peptidic small molecules for exogenous control of cellular processes. The modular structure of huezole provides a framework for designing a class of organelle-emulating small molecules.

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