RESUMO
Hormones and neurotransmitters are essential to homeostasis, and their disruptions are connected to diseases ranging from cancer to anxiety. The differential reactivation of endobiotic glucuronides by gut microbial ß-glucuronidase (GUS) enzymes may influence interindividual differences in the onset and treatment of disease. Using multi-omic, in vitro, and in vivo approaches, we show that germ-free mice have reduced levels of active endobiotics and that distinct gut microbial Loop 1 and FMN GUS enzymes drive hormone and neurotransmitter reactivation. We demonstrate that a range of FDA-approved drugs prevent this reactivation by intercepting the catalytic cycle of the enzymes in a conserved fashion. Finally, we find that inhibiting GUS in conventional mice reduces free serotonin and increases its inactive glucuronide in the serum and intestines. Our results illuminate the indispensability of gut microbial enzymes in sustaining endobiotic homeostasis and indicate that therapeutic disruptions of this metabolism promote interindividual response variabilities.
Assuntos
Microbioma Gastrointestinal , Glucuronidase , Homeostase , Animais , Microbioma Gastrointestinal/efeitos dos fármacos , Camundongos , Glucuronidase/metabolismo , Camundongos Endogâmicos C57BL , Serotonina/metabolismo , Glucuronídeos/metabolismo , Humanos , Intestinos/microbiologia , Masculino , Vida Livre de GermesRESUMO
Cyclic peptides are poised to target historically difficult to drug intracellular protein-protein interactions, however, their general cell impermeability poses a challenge for characterizing function. Recent advances in microfluidics have enabled permeabilization of the cytoplasmic membrane by physical cell deformation (i.e., mechanoporation), resulting in intracellular delivery of impermeable macromolecules in vector- and electrophoretic-free approaches. However, the number of payloads (e.g., peptides) and/or concentrations delivered via microfluidic mechanoporation is limited by having to pre-mix cells and payloads, a manually intensive process. In this work, we show that cells are momentarily permeable (t 1/2 = 1.1-2.8 min) after microfluidic vortex shedding (µVS) and that lower molecular weight macromolecules can be cytosolically delivered upon immediate exposure after cells are processed/permeabilized. To increase the ability to screen peptides, we built a system, dispensing-microfluidic vortex shedding (DµVS), that integrates a µVS chip with inline microplate-based dispensing. To do so, we synced an electronic pressure regulator, flow sensor, on/off dispense valve, and an x-y motion platform in a software-driven feedback loop. Using this system, we were able to deliver low microliter-scale volumes of transiently mechanoporated cells to hundreds of wells on microtiter plates in just several minutes (e.g., 96-well plate filled in <2.5 min). We validated the delivery of an impermeable peptide directed at MDM2, a negative regulator of the tumor suppressor p53, using a click chemistry- and NanoBRET-based cell permeability assay in 96-well format, with robust delivery across the full plate. Furthermore, we demonstrated that DµVS could be used to identify functional, low micromolar, cellular activity of otherwise cell-inactive MDM2-binding peptides using a p53 reporter cell assay in 96- and 384-well format. Overall, DµVS can be combined with downstream cell assays to investigate intracellular target engagement in a high-throughput manner, both for improving structure-activity relationship efforts and for early proof-of-biology of non-optimized peptide (or potentially other macromolecular) tools.
RESUMO
Target identification involves deconvoluting the protein target of a pharmacologically active, small-molecule ligand, a process that is critical for early drug discovery yet technically challenging. Photoaffinity labelling strategies have become the benchmark for small-molecule target deconvolution, but covalent protein capture requires the use of high-energy ultraviolet light, which can complicate downstream target identification. Thus, there is a strong demand for alternative technologies that allow for controlled activation of chemical probes to covalently label their protein target. Here we introduce an electroaffinity labelling platform that leverages the use of a small, redox-active diazetidinone functional group to enable chemoproteomic-based target identification of pharmacophores within live cell environments. The underlying discovery to enable this platform is that the diazetidinone can be electrochemically oxidized to reveal a reactive intermediate useful for covalent modification of proteins. This work demonstrates the electrochemical platform to be a functional tool for drug-target identification.
Assuntos
Descoberta de Drogas , Proteínas , Proteínas/metabolismo , Marcadores de Fotoafinidade/química , Ligantes , FarmacóforoRESUMO
The growing appreciation of immune cell-cell interactions within disease environments has led to extensive efforts to develop immunotherapies. However, characterizing complex cell-cell interfaces in high resolution remains challenging. Thus, technologies leveraging therapeutic-based modalities to profile intercellular environments offer opportunities to study cell-cell interactions with molecular-level insight. We introduce photocatalytic cell tagging (PhoTag) for interrogating cell-cell interactions using single-domain antibodies (VHHs) conjugated to photoactivatable flavin-based cofactors. Following irradiation with visible light, the flavin photocatalyst generates phenoxy radical tags for targeted labeling. Using this technology, we demonstrate selective synaptic labeling across the PD-1/PD-L1 axis in antigen-presenting cell-T cell systems. In combination with multiomics single-cell sequencing, we monitored interactions between peripheral blood mononuclear cells and Raji PD-L1 B cells, revealing differences in transient interactions with specific T cell subtypes. The utility of PhoTag in capturing cell-cell interactions will enable detailed profiling of intercellular communication across different biological systems.
Assuntos
Antígeno B7-H1 , Leucócitos Mononucleares , Comunicação Celular , Flavinas , ImunoterapiaRESUMO
Despite the growing use of visible-light photochemistry in both chemistry and biology, no general low-heat photoreactor for use across these different disciplines exists. Herein, we describe the design and use of a standardized photoreactor for visible-light-driven activation and photocatalytic chemical transformations. Using this single benchtop photoreactor, we performed photoredox reactions across multiple visible light wavelengths, a high-throughput photocatalytic cross-coupling reaction, and inâ vitro labeling of proteins and live cells. Given the success of this reactor in all tested applications, we envision that this multi-use photoreactor will be widely used in biology, chemical biology, and medicinal chemistry settings.
Assuntos
Biotina/análise , Luz , Fotobiorreatores , Tiramina/química , Catálise , Linhagem Celular Tumoral , Desenho de Equipamento , Humanos , Estrutura Molecular , Processos Fotoquímicos , Tiramina/análogos & derivados , Tiramina/síntese químicaRESUMO
It is increasingly clear that interindividual variability in human gut microbial composition contributes to differential drug responses. For example, gastrointestinal (GI) toxicity is not observed in all patients treated with the anticancer drug irinotecan, and it has been suggested that this variability is a result of differences in the types and levels of gut bacterial ß-glucuronidases (GUSs). GUS enzymes promote drug toxicity by hydrolyzing the inactive drug-glucuronide conjugate back to the active drug, which damages the GI epithelium. Proteomics-based identification of the exact GUS enzymes responsible for drug reactivation from the complexity of the human microbiota has not been accomplished, however. Here, we discover the specific bacterial GUS enzymes that generate SN-38, the active and toxic metabolite of irinotecan, from human fecal samples using a unique activity-based protein profiling (ABPP) platform. We identify and quantify gut bacterial GUS enzymes from human feces with an ABPP-enabled proteomics pipeline and then integrate this information with ex vivo kinetics to pinpoint the specific GUS enzymes responsible for SN-38 reactivation. Furthermore, the same approach also reveals the molecular basis for differential gut bacterial GUS inhibition observed between human fecal samples. Taken together, this work provides an unprecedented technical and bioinformatics pipeline to discover the microbial enzymes responsible for specific reactions from the complexity of human feces. Identifying such microbial enzymes may lead to precision biomarkers and novel drug targets to advance the promise of personalized medicine.
Assuntos
Proteínas de Bactérias/metabolismo , Cicloexanóis/química , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/metabolismo , Inibidores Enzimáticos/química , Microbioma Gastrointestinal/fisiologia , Glucuronidase/metabolismo , Irinotecano/química , Animais , Biomarcadores/metabolismo , Biologia Computacional , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/microbiologia , Inibidores Enzimáticos/metabolismo , Fezes/química , Feminino , Glucuronídeos/metabolismo , Humanos , Hidrólise , Irinotecano/metabolismo , Cinética , Masculino , Metaboloma , Camundongos , Modelos Moleculares , Medicina de Precisão , Ligação Proteica , Conformação ProteicaRESUMO
Gut microbial ß-glucuronidase (GUS) enzymes have been suggested to be involved in the estrobolome, the collection of microbial reactions involving estrogens. Furthermore, bacterial GUS enzymes within the gastrointestinal tract have been postulated to be a contributing factor in hormone-driven cancers. However, to date, there has been no experimental evidence to support these hypotheses. Here we provide the first in vitro analysis of the ability of 35 human gut microbial GUS enzymes to reactivate two distinct estrogen glucuronides, estrone-3-glucuronide and estradiol-17-glucuronide, to estrone and estradiol, respectively. We show that certain members within the Loop 1, mini-Loop 1, and FMN-binding classes of gut microbial GUS enzymes can reactivate estrogens from their inactive glucuronides. We provide molecular details of key interactions that facilitate these catalytic processes and present the structures of two novel human gut microbial GUS enzymes related to the estrobolome. Further, we demonstrate that estrogen reactivation by Loop 1 bacterial GUS enzymes can be inhibited both in purified enzymes and in fecal preparations of mixed murine fecal microbiota. Finally, however, despite these in vitro and ex vivo data, we show that a Loop 1 GUS-specific inhibitor is not capable of reducing the development of tumors in the PyMT mouse model of breast cancer. These findings validate that gut microbial GUS enzymes participate in the estrobolome but also suggest that the estrobolome is a multidimensional set of processes on-going within the mammalian gastrointestinal tract that likely involves many enzymes, including several distinct types of GUS proteins.
Assuntos
Estrogênios/metabolismo , Glucuronidase/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Estrona/metabolismo , Feminino , Microbioma Gastrointestinal/fisiologia , Glucuronidase/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mutagênese Sítio-DirigidaRESUMO
Histone deacetylases (HDACs) regulate diverse cellular processes, and are promising targets for a number of diseases. Here we describe the design and utilization of a largazole-based chemical probe to quantitatively measure the intracellular occupancy of HDAC1 and HDAC2 by dacinostat. Surprisingly, the probe was unable to enrich HDAC3 despite its nanomolar potency in a biochemical assay, further proving the necessity of cell-based target occupancy assays to understand compound potency in physiologically-relevant settings. This occupancy assay has the potential to aid the development of novel HDAC1/2 inhibitors in drug discovery.
RESUMO
BACKGROUND: Exacerbations of asthma and COPD are triggered by rhinoviruses. Uncontrolled inflammatory pathways, pathogenic bacterial burden and impaired antiviral immunity are thought to be important factors in disease severity and duration. Macrolides including azithromycin are often used to treat the above diseases, but exhibit variable levels of efficacy. Inhaled corticosteroids are also readily used in treatment, but may lack specificity. Ideally, new treatment alternatives should suppress unwanted inflammation, but spare beneficial antiviral immunity. METHODS: In the present study, we screened 225 novel macrolides and tested them for enhanced antiviral activity against rhinovirus, as well as anti-inflammatory activity and activity against Gram-positive and Gram-negative bacteria. Primary bronchial epithelial cells were grown from 10 asthmatic individuals and the effects of macrolides on rhinovirus replication were also examined. Another 30 structurally similar macrolides were also examined. RESULTS: The oleandomycin derivative Mac5, compared with azithromycin, showed superior induction (up to 5-fold, EC50â=â5-11 µM) of rhinovirus-induced type I IFNß, type III IFNλ1 and type III IFNλ2/3 mRNA and the IFN-stimulated genes viperin and MxA, yet had no effect on IL-6 and IL-8 mRNA. Mac5 also suppressed rhinovirus replication at 48 h, proving antiviral activity. Mac5 showed antibacterial activity against Gram-positive Streptococcus pneumoniae; however, it did not have any antibacterial properties compared with azithromycin when used against Gram-negative Escherichia coli (as a model organism) and also the respiratory pathogens Pseudomonas aeruginosa and non-typeable Haemophilus influenzae. Further non-toxic Mac5 derivatives were identified with various anti-inflammatory, antiviral and antibacterial activities. CONCLUSIONS: The data support the idea that macrolides have antiviral properties through a mechanism that is yet to be ascertained. We also provide evidence that macrolides can be developed with anti-inflammatory, antibacterial and antiviral activity and show surprising versatility depending on the clinical need.
Assuntos
Antibacterianos/farmacologia , Anti-Inflamatórios não Esteroides/farmacologia , Antivirais/química , Antivirais/farmacologia , Descoberta de Drogas , Interferons/imunologia , Macrolídeos/farmacologia , Antibacterianos/química , Antibacterianos/isolamento & purificação , Antibacterianos/uso terapêutico , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/isolamento & purificação , Anti-Inflamatórios não Esteroides/uso terapêutico , Antivirais/isolamento & purificação , Antivirais/uso terapêutico , Asma/tratamento farmacológico , Brônquios/citologia , Brônquios/efeitos dos fármacos , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Células Epiteliais/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Haemophilus influenzae/efeitos dos fármacos , Humanos , Interferon beta/imunologia , Interferons/biossíntese , Interleucina-6/imunologia , Interleucina-6/metabolismo , Interleucina-8/imunologia , Interleucina-8/metabolismo , Macrolídeos/química , Macrolídeos/uso terapêutico , Proteínas de Resistência a Myxovirus/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-CH , Proteínas/genética , Pseudomonas aeruginosa/efeitos dos fármacos , Rhinovirus/efeitos dos fármacos , Replicação Viral/efeitos dos fármacosRESUMO
Peptide macrocycles represent a chemical space where the best of biological tools can synergize with the best of chemical approaches in the quest for leads against undruggable targets. Peptide macrocycles offer some key advantages in both lead discovery and lead optimization phases of drug discovery when compared to natural product and synthetic macrocycles. In addition, they are uniquely positioned to capitalize on the therapeutic potential of peptides because cyclization can help drive selectivity, potency and overcome the common limitations of metabolic instability of peptides.
Assuntos
Descoberta de Drogas , Compostos Macrocíclicos/farmacologia , Peptídeos/farmacologia , Ciclização , Humanos , Compostos Macrocíclicos/síntese química , Compostos Macrocíclicos/química , Compostos Macrocíclicos/uso terapêutico , Peptídeos/síntese química , Peptídeos/química , Peptídeos/uso terapêutico , Estabilidade ProteicaRESUMO
Kinases are principal components of signal transduction pathways and the focus of intense basic and drug discovery research. Irreversible inhibitors that covalently modify non-catalytic cysteines in kinase active sites have emerged as valuable probes and approved drugs. Many protein classes, however, have functional cysteines, and therefore understanding the proteome-wide selectivity of covalent kinase inhibitors is imperative. Here, we accomplish this objective using activity-based protein profiling coupled with quantitative MS to globally map the targets, both specific and nonspecific, of covalent kinase inhibitors in human cells. Many of the specific off-targets represent nonkinase proteins that, notably, have conserved active site cysteines. We define windows of selectivity for covalent kinase inhibitors and show that, when these windows are exceeded, rampant proteome-wide reactivity and kinase target-independent cell death conjointly occur. Our findings, taken together, provide an experimental road map to illuminate opportunities and surmount challenges for the development of covalent kinase inhibitors.
Assuntos
Inibidores de Proteínas Quinases/farmacologia , Proteoma/genética , Adenina/análogos & derivados , Tirosina Quinase da Agamaglobulinemia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisteína/química , Genes erbB-1/genética , Humanos , Cinética , Piperidinas , Proteínas Quinases/metabolismo , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirazóis/farmacologia , Pirimidinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
A series of 4-substituted pyrimido[4,5-d]azepines that are potent, selective 5-HT2C receptor partial agonists is described. A rational medicinal chemistry design strategy to deliver CNS penetration coupled with SAR-based optimization of selectivity and agonist potency provided compounds with the desired balance of preclinical properties. Lead compounds 17 (PF-4479745) and 18 (PF-4522654) displayed robust pharmacology in a preclinical canine model of stress urinary incontinence (SUI) and no measurable functional agonism at the key selectivity targets 5-HT2A and 5-HT2B in relevant tissue-based assay systems. Utilizing recent advances in the structural biology of GPCRs, homology modeling has been carried out to rationalize binding and agonist efficacy of these compounds.
Assuntos
Azepinas/química , Fármacos do Sistema Nervoso Central/química , Pirimidinas/química , Receptor 5-HT2A de Serotonina/metabolismo , Receptor 5-HT2B de Serotonina/metabolismo , Receptor 5-HT2C de Serotonina/metabolismo , Agonistas do Receptor 5-HT2 de Serotonina/química , Animais , Azepinas/síntese química , Azepinas/farmacologia , Barreira Hematoencefálica/metabolismo , Células CHO , Fármacos do Sistema Nervoso Central/síntese química , Fármacos do Sistema Nervoso Central/farmacologia , Cricetulus , Cães , Desenho de Fármacos , Humanos , Células Madin Darby de Rim Canino , Permeabilidade , Pirimidinas/síntese química , Pirimidinas/farmacologia , Agonistas do Receptor 5-HT2 de Serotonina/síntese química , Agonistas do Receptor 5-HT2 de Serotonina/farmacologia , Relação Estrutura-Atividade , Incontinência Urinária por Estresse/tratamento farmacológicoRESUMO
In ongoing studies towards novel hepatitisâ C virus (HCV) therapeutics, inhibitors of nonstructural protein 5A (NS5A) were evaluated. Specifically, starting from previously reported lead compounds, peripheral substitution patterns of a series of biaryl-linked pyrrolidine NS5A replication complex inhibitors were probed and structure-activity relationships were elucidated. Using molecular modelling and a supercritical fluid chromatographic (SFC) technique, intramolecular H-bonding and peripheral functional group topology were evaluated as key determinants of activity and membrane permeability. The novel compounds exhibited retained potency as compared with the lead compounds, and also showed promising results against a panel of resistance viruses. Together, the results of the study take us a step closer towards understanding the potency of daclatasvir, a clinical candidate upon which the compounds were based, and to designing improved analogues as second-generation antiviral agents targeting NS5A.
Assuntos
Inibidores de Proteases/química , Proteínas não Estruturais Virais/antagonistas & inibidores , Animais , Antivirais/síntese química , Antivirais/química , Antivirais/farmacologia , Linhagem Celular , Permeabilidade da Membrana Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cães , Avaliação Pré-Clínica de Medicamentos , Farmacorresistência Viral , Hepacivirus/metabolismo , Humanos , Ligação de Hidrogênio , Inibidores de Proteases/síntese química , Inibidores de Proteases/farmacologia , Ratos , Relação Estrutura-Atividade , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/efeitos dos fármacosRESUMO
Small peptidic kappa agonists were covalently linked to the reactive lysine of the CovX antibody to create compounds having potent activity at the kappa receptor with greatly extended half-life when compared to the parent peptide as exemplified by compound 20.
Assuntos
Analgésicos/síntese química , Imunoconjugados/química , Peptídeos Opioides/síntese química , Receptores Opioides kappa/agonistas , Bibliotecas de Moléculas Pequenas/síntese química , Analgésicos/farmacologia , Animais , Anticorpos/química , Arrestinas/metabolismo , Azetidinas/química , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos , Meia-Vida , Humanos , Imunoconjugados/farmacologia , Ligantes , Peptídeos Opioides/farmacologia , Transporte Proteico/efeitos dos fármacos , Ratos , Receptores Opioides kappa/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-Atividade , beta-ArrestinasRESUMO
3-Azabicyclo[3.1.0]hexane compounds were designed as novel achiral µ opioid receptor ligands for the treatment of pruritus in dogs. In this paper, we describe the SAR of this class of opioid ligand, highlighting changes to the lead structure which led to compounds having picomolar binding affinity, selective for the µ receptor over δ and κ subtypes. Some subtleties of functional activity will also be described.
Assuntos
Antipruriginosos/síntese química , Compostos Bicíclicos Heterocíclicos com Pontes/síntese química , Hexanos/síntese química , Prurido/tratamento farmacológico , Receptores Opioides mu/antagonistas & inibidores , Animais , Antipruriginosos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Cães , Cobaias , Hexanos/farmacologia , Humanos , Técnicas In Vitro , Cinética , Ligantes , Prurido/metabolismo , Receptores Opioides delta/antagonistas & inibidores , Receptores Opioides delta/metabolismo , Receptores Opioides kappa/antagonistas & inibidores , Receptores Opioides kappa/metabolismo , Receptores Opioides mu/metabolismo , Relação Estrutura-AtividadeRESUMO
A heterotrifunctional template was developed that utilizes thiol-maleimide and click chemistries (both copper-free and copper-mediated) to effect sequential biomolecule conjugations in a one-pot process. The breadth of compatible substrates was illustrated through highly efficient conjugations of protein, peptide, sugar, lipid, fluoroalkane, biotin and fluorophore molecules. This template should be useful for the creation of chemically-enhanced/enabled biotherapeutics, especially through the expression of discontinuous (and heterogeneous) epitopes.
Assuntos
Química Click , Proteínas/química , Animais , Bovinos , Modelos Moleculares , Conformação Proteica , Soroalbumina Bovina/química , EstereoisomerismoRESUMO
The V1a receptor has emerged as an attractive target for a range of indications including Raynaud's disease and dysmenorrhoea. As part of an effort to discover a new class of orally active V1a antagonist, we optimised a highly lipophilic, metabolically unstable lead into a range of potent, selective and metabolically stable V1a antagonists. In this communication, we demonstrate the series-dependent effect of limiting the number of rotatable bonds in order to decrease Cytochrome P450-mediated metabolism. This effort culminated in the discovery of PF-184563, a novel, selective V1a antagonist with excellent in vitro and in vivo properties.
Assuntos
Antagonistas dos Receptores de Hormônios Antidiuréticos , Benzodiazepinas/síntese química , Benzodiazepinas/farmacologia , Sistema Enzimático do Citocromo P-450/metabolismo , Desenho de Fármacos , Descoberta de Drogas , Dismenorreia/tratamento farmacológico , Antagonistas de Hormônios/síntese química , Antagonistas de Hormônios/farmacologia , Triazóis/síntese química , Triazóis/farmacologia , Benzodiazepinas/química , Benzodiazepinas/metabolismo , Estabilidade de Medicamentos , Feminino , Antagonistas de Hormônios/química , Antagonistas de Hormônios/metabolismo , Humanos , Microssomos/fisiologia , Estrutura Molecular , Triazóis/química , Triazóis/metabolismoRESUMO
A series of p-hydroxybenzenesulphonamides ERß receptor agonists were discovered and several compounds identified had excellent selectivity over the related ERα receptor. One of these, compound 11, had an interesting binding conformation determined by X-ray and represents an excellent starting point in the quest for further selective ERß agonists.
Assuntos
Descoberta de Drogas , Receptor beta de Estrogênio/agonistas , Sulfonamidas/química , Sulfonamidas/farmacologia , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Humanos , Ligantes , Modelos Químicos , Ligação Proteica , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/metabolismoRESUMO
A series of acidic triazoles with activity as soluble guanylate cyclase stimulators is described. Incorporation of the CF(3) triazole improved the overall physicochemical and drug-like properties of the molecule and is exemplified by compound 25.
Assuntos
Ácidos/química , Ativadores de Enzimas/farmacologia , Guanilato Ciclase/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Triazóis/farmacologia , Guanilil Ciclase Solúvel , Triazóis/químicaRESUMO
Synthetic peptides that specifically bind nuclear hormone receptors offer an alternative approach to small molecules for the modulation of receptor signaling and subsequent gene expression. Here we describe the design of a series of novel stapled peptides that bind the coactivator peptide site of estrogen receptors. Using a number of biophysical techniques, including crystal structure analysis of receptor-stapled peptide complexes, we describe in detail the molecular interactions and demonstrate that all-hydrocarbon staples modulate molecular recognition events. The findings have implications for the design of stapled peptides in general.