RESUMEN
Protein folding is assisted by molecular chaperones that bind nascent polypeptides during mRNA translation. Several structurally distinct classes of chaperones promote de novo folding, suggesting that their activities are coordinated at the ribosome. We used biochemical reconstitution and structural proteomics to explore the molecular basis for cotranslational chaperone action in bacteria. We found that chaperone binding is disfavored close to the ribosome, allowing folding to precede chaperone recruitment. Trigger factor recognizes compact folding intermediates that expose an extensive unfolded surface, and dictates DnaJ access to nascent chains. DnaJ uses a large surface to bind structurally diverse intermediates and recruits DnaK to sequence-diverse solvent-accessible sites. Neither Trigger factor, DnaJ, nor DnaK destabilize cotranslational folding intermediates. Instead, the chaperones collaborate to protect incipient structure in the nascent polypeptide well beyond the ribosome exit tunnel. Our findings show how the chaperone network selects and modulates cotranslational folding intermediates.
Asunto(s)
Proteínas de Escherichia coli , Escherichia coli , Proteínas del Choque Térmico HSP40 , Proteínas HSP70 de Choque Térmico , Biosíntesis de Proteínas , Pliegue de Proteína , Ribosomas , Ribosomas/metabolismo , Ribosomas/genética , Proteínas de Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/química , Proteínas HSP70 de Choque Térmico/metabolismo , Proteínas HSP70 de Choque Térmico/genética , Proteínas del Choque Térmico HSP40/metabolismo , Proteínas del Choque Térmico HSP40/genética , Escherichia coli/metabolismo , Escherichia coli/genética , Unión Proteica , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genética , Modelos Moleculares , Conformación Proteica , Isomerasa de PeptidilprolilRESUMEN
PROTACs have recently emerged as a novel paradigm in drug discovery. They can hijack existing biological machinery to selectively degrade proteins of interest, in a catalytic fashion. Here we describe the design, optimisation and biological activity of a set of novel PROTACs targeting the Janus kinase family (JAK1, JAK2, JAK3 and TYK2) of proximal membrane-bound proteins. The JAK family proteins display membrane localisation by virtue of their association with cytoplasmic tails of cytokine receptors, and there are no reports of a successful PROTAC strategy being deployed against this class of proteins. JAK PROTACs from two distinct JAK chemotypes were designed, optimising the physicochemical properties for each template to enhance cell permeation. These PROTACs are capable of inducing JAK1 and JAK2 degradation, demonstrating an extension of the PROTAC methodology to an unprecedented class of protein targets. A number of known ligase binders were explored, and it was found that PROTACs bearing an inhibitor of apoptosis protein (IAP) ligand induced significantly more JAK degradation over Von Hippel-Lindau (VHL) and Cereblon (CRBN) PROTACs. In addition, the mechanism of action of the JAK PROTACs was elucidated, and it was confirmed that JAK degradation was both IAP- and proteasome-dependent.
Asunto(s)
Quinasas Janus/antagonistas & inhibidores , Proteolisis/efectos de los fármacos , Pirimidinas/farmacología , Quinoxalinas/farmacología , Factores de Transcripción STAT/antagonistas & inhibidores , Ubiquitina/antagonistas & inhibidores , Relación Dosis-Respuesta a Droga , Diseño de Fármacos , Humanos , Quinasas Janus/metabolismo , Ligandos , Estructura Molecular , Pirimidinas/síntesis química , Pirimidinas/química , Quinoxalinas/síntesis química , Quinoxalinas/química , Factores de Transcripción STAT/metabolismo , Relación Estructura-Actividad , Células THP-1 , Ubiquitina/metabolismoRESUMEN
Palladium-catalysed coupling reactions based on a novel and easy-to-synthesise difluorinated organotrifluoroborate were used to assemble precursors to 6π-electrocyclisations of three different types. Electrocyclisations took place at temperatures between 90 and 240 °C, depending on the central component of the π-system; nonaromatic trienes were most reactive, but even systems that required the temporary dearomatisation of two arenyl subunits underwent electrocyclisation, albeit at elevated temperatures. Photochemical conditions were effective for these more demanding reactions. The package of methods delivered a structurally diverse set of fluorinated arenes, spanning a 20â kcal mol(-1) range of reactivity, by a flexible route.
RESUMEN
OBJECTIVES: (S)-Leucoxine, isolated from the Colombian Lauraceae tree Rhodostemonodaphne crenaticupula Madriñan, was found to inhibit the growth of Mycobacterium tuberculosis H37Rv. A biomimetic approach for the chemical synthesis of a wide array of 1-substituted tetrahydroisoquinolines was undertaken with the aim of elucidating a common pharmacophore for these compounds with novel mode(s) of anti-TB action. METHODS: Biomimetic Pictet-Spengler or Bischler-Napieralski synthetic routes were employed followed by an evaluation of the biological activity of the synthesized compounds. RESULTS: In this work, the synthesized tetrahydroisoquinolines were found to inhibit the growth of M. tuberculosis H37Rv and affect its whole-cell phenotype as well as the activity of the ATP-dependent MurE ligase, a key enzyme involved in the early stage of cell wall peptidoglycan biosynthesis. CONCLUSIONS: As the correlation between the MIC and the half-inhibitory enzymatic concentration was not particularly strong, there is a credible possibility that these compounds have pleiotropic mechanism(s) of action in M. tuberculosis.
Asunto(s)
Antituberculosos/farmacología , Proteínas Bacterianas/efectos adversos , Inhibidores Enzimáticos/farmacología , Mycobacterium tuberculosis/efectos de los fármacos , Péptido Sintasas/efectos adversos , Tetrahidroisoquinolinas/farmacología , Antituberculosos/síntesis química , Pruebas de Sensibilidad Microbiana , Mycobacterium tuberculosis/enzimología , Mycobacterium tuberculosis/crecimiento & desarrollo , Tetrahidroisoquinolinas/síntesis químicaRESUMEN
Boronic acid solution speciation can be controlled during the Suzuki-Miyaura cross-coupling of haloaryl N-methyliminodiacetic acid (MIDA) boronic esters to enable the formal homologation of boronic acid derivatives. The reaction is contingent upon control of the basic biphase and is thermodynamically driven: temperature control provides highly chemoselective access to either BMIDA adducts at room temperature or boronic acid pinacol ester (BPin) products at elevated temperature. Control experiments and solubility analyses have provided some insight into the mechanistic operation of the formal homologation process.
Asunto(s)
Ácidos Borónicos/química , Ésteres/química , Catálisis , Estructura MolecularRESUMEN
4H-Quinolizin-4-ones are a unique class of heterocycle with valuable physicochemical properties and which are emerging as key pharmacophores for a range of biological targets. A tandem Horner-Wadsworth-Emmons olefination/cyclisation method has been developed to allow facile access to substituted 4H-quinolizin-4-ones encoded with a range of functional groups.
Asunto(s)
Alquenos/química , Quinolizinas/síntesis química , Ciclización , Estructura Molecular , Quinolizinas/químicaRESUMEN
A recently developed method for the near-ambient generation of difluorovinylzinc reagents has facilitated the preparation of 1-(N,N-diethylcarbamoyloxy)-2,2-difluoro-1-iodoethene and 2,2-difluoro-1-iodo-1-(2'-methoxyethoxymethoxy)ethene. The utility of these reagents has been investigated in Suzuki-Miyaura couplings with a range of potassium trifluoroborate coupling partners, with the scope of successful couplings proving wide. Deiodinated species appeared as significant side products, but a solvent change from i-PrOH to t-BuOH suppressed the pathway to these species and improved coupling yields.
Asunto(s)
Hidrocarburos Fluorados/síntesis química , Hidrocarburos Yodados/síntesis química , Temperatura , Hidrocarburos Fluorados/química , Hidrocarburos Yodados/química , Estructura Molecular , EstereoisomerismoRESUMEN
A major limitation of pulmonary delivery is that drugs can exhibit suboptimal pharmacokinetic profiles resulting from rapid elimination from the pulmonary tissue. This can lead to systemic side effects and a short duration of action. A series of dibasic dipeptides attached to the poorly lung-retentive muscarinic M3 receptor antagonist piperidin-4-yl 2-hydroxy-2,2-diphenylacetate (1) through a pH-sensitive-linking group have been evaluated. Extensive optimization resulted in 1-(((R)-2-((S)-2,6-diaminohexanamido)-3,3-dimethylbutanoyl)oxy)ethyl 4-(2-hydroxy-2,2-diphenylacetoxy)piperidine-1-carboxylate (23), which combined very good in vitro stability and very high rat lung binding. Compound 23 progressed to pharmacokinetic studies in rats, where, at 24 h post dosing in the rat lung, the total lung concentration of 23 was 31.2 µM. In addition, high levels of liberated drug 1 were still detected locally, demonstrating the benefit of this novel prodrug approach for increasing the apparent pharmacokinetic half-life of drugs in the lungs following pulmonary dosing.
Asunto(s)
Profármacos , Animales , Semivida , Pulmón , Antagonistas Muscarínicos/farmacología , Profármacos/química , RatasRESUMEN
Difluoroalkenylzinc reagents prepared from 1-(2'-methoxy-ethoxymethoxy)-2,2,2-trifluoroethane and 1-(N,N-diethylcarbamoyloxy)-2,2,2-trifluoroethane at ice bath temperatures underwent Negishi coupling with a range of aryl halides in a convenient one pot procedure. While significant differences between the enol acetal and carbamate reagents were revealed, the Negishi protocol compared very favourably with alternative coupling procedures in terms of overall yields from trifluoroethanol.
Asunto(s)
Hidrocarburos Fluorados/química , Compuestos Organometálicos/síntesis química , Temperatura , Trifluoroetanol/química , Estructura Molecular , Compuestos Organometálicos/química , EstereoisomerismoRESUMEN
Associated with their roles as targets for tumour promoters, there has been a long-standing interest in how members of the protein kinase C (PKC) family act to modulate cell growth and division. This has generated a great deal of observational data, but has for the most part not afforded clear mechanistic insights into the control mechanisms at play. Here, we review the roles of PKCε in protecting transformed cells from non-disjunction. In this particular cell cycle context, there is a growing understanding of the pathways involved, affording biomarker and interventional insights and opportunities.
Asunto(s)
Genómica , Neoplasias/genética , Proteína Quinasa C-epsilon/metabolismo , Proliferación Celular , Humanos , Neoplasias/enzimología , Neoplasias/patologíaRESUMEN
Enantiopure bromonium ions may be generated from enantiopure bromohydrins and derivatives, they can be trapped with an in situ nucleophile to give enantiomerically pure products.
Asunto(s)
Alcoholes/química , Bromuros/química , Bromuros/síntesis química , EstereoisomerismoRESUMEN
4-(Pyrimidin-4-yl)morpholines are privileged pharmacophores for PI3K and PIKKs inhibition by virtue of the morpholine oxygen, both forming the key hydrogen bonding interaction and conveying selectivity over the broader kinome. Key to the morpholine utility as a kinase hinge binder is its ability to adopt a coplanar conformation with an adjacent aromatic core favored by the morpholine nitrogen nonbonding pair of electrons interacting with the electron deficient pyrimidine π-system. Few selective morpholine replacements have been identified to date. Herein we describe the discovery of a potent non-nitrogen containing morpholine isostere with the ability to mimic this conformation and its application in a potent selective dual inhibitor of mTORC1 and mTORC2 (29b).