RESUMEN
A totally new approach in the synthesis of mixed polymer brushes tethered on polyamide (PA) surfaces is presented herein. As a proof of concept, two types of homopolymers were synthesized in sequential surface-initiated atom transfer radical polymerization (SI-ATRP) reactions: poly(methyl methacrylate)/poly((2-dimethylamino)ethyl methacrylate) and polystyrene /poly((2-dimethylamino)ethyl methacrylate). The ATRP initiator was immobilized on the surface through PA chain-end groups in two subsequent steps, separated by homo-polymerizations. The amount of the PA chains' end groups available on the modified surface was tuned by the thermal rearrangement of the surface.
RESUMEN
Protein kinases are widely recognized as important therapeutic targets due to their involvement in signal transduction pathways. These pathways are tightly controlled and regulated, notably by the ability of kinases to selectively phosphorylate a defined set of substrates. A wide variety of disorders can arise as a consequence of abnormal kinase-mediated phosphorylation and numerous kinase inhibitors have earned their place as key components of the modern pharmacopeia. Although "traditional" kinase inhibitors typically act by preventing the interaction between the kinase and ATP, thus stopping substrate phosphorylation, an alternative approach consists in disrupting the protein-protein interaction between the kinase and its downstream partners. In order to facilitate the identification of potential chemical starting points for substrate-site inhibition approaches, we desired to investigate the application of Substrate Activity Screening to kinases. We herein report a proof-of-concept study demonstrating, on a model tyrosine kinase, that the key requirements of this methodology can be met. Namely, using peptides as model substrates, we show that a simple ADP-accumulation assay can be used to monitor substrate efficiency and that efficiency can be optimized in a modular manner. More importantly, we demonstrate that structure-efficiency relationships translate into structure-activity relationships upon conversion of the substrates into inhibitors.
Asunto(s)
Péptidos/química , Inhibidores de Proteínas Quinasas/química , Proteínas Tirosina Quinasas Receptoras/química , Adenosina Trifosfato/química , Ensayos Analíticos de Alto Rendimiento , Humanos , Cinética , Péptidos/antagonistas & inhibidores , Unión Proteica , Inhibidores de Proteínas Quinasas/síntesis química , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Proteínas Recombinantes de Fusión/antagonistas & inhibidores , Proteínas Recombinantes de Fusión/química , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
Protein kinases are widely recognized as important therapeutic targets due to their involvement in signal transduction pathways. These pathways are tightly controlled and regulated, notably by the ability of kinases to selectively phosphorylate a defined set of substrates. As part of a study on the substrate requirements of Insulin-like Growth Factor 1 Receptor (IGF-1R) and Insulin Receptor (InsR), we evaluated and applied a universal assay system able to monitor the phosphorylation of unlabelled peptides of any length in real time. In contrast to already reported profiling methodologies, we were able to assess the k(cat)/K(M) ratio of peptides as short as tetramers. Notably, we were able to identify an efficient pentamer substrate that exhibited kinetic properties close to those of a 250-amino acid protein derived from IRS-1, a natural substrate of IGF-1R and InsR.
Asunto(s)
Sondas Moleculares/química , Péptidos/química , Receptor IGF Tipo 1/química , Secuencia de Aminoácidos , Cristalografía por Rayos X , Modelos Moleculares , Sondas Moleculares/genética , Datos de Secuencia Molecular , Péptidos/genética , Fosforilación , Unión Proteica , Receptor de Insulina/química , Receptor de Insulina/genética , Especificidad por SustratoRESUMEN
The vitamin E analogues (2R,4'R,8'R)-nor-α-tocopherol (94 % de) and (2RS,4'R,8'R)-nor-α-tocopherol have been synthesized from (all R)-hexahydrofarnesol and phytol, respectively. According to in vitro experiments with murine macrophages nor-α-tocopherol is an anti-inflammatory compound more potent than α-tocopherol.
Asunto(s)
Antioxidantes/síntesis química , Antioxidantes/farmacología , alfa-Tocoferol/síntesis química , alfa-Tocoferol/farmacología , Animales , Antioxidantes/química , Línea Celular , Citocinas/metabolismo , Inflamación/genética , Macrófagos/efectos de los fármacos , Macrófagos/enzimología , Macrófagos/metabolismo , Ratones , Modelos Moleculares , Conformación Molecular , Óxido Nítrico Sintasa de Tipo II/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Estereoisomerismo , alfa-Tocoferol/análogos & derivados , alfa-Tocoferol/químicaRESUMEN
A diastereoselective synthesis of alpha-tocopherol 1 (93% de) was achieved via two key steps, (i) a highly diastereoselective Shi epoxidation of a trisubstituted alkene and (ii) an acid supported, "anti-Baldwin" epoxide ring opening under inversion of configuration leading to the 6-membered chromanol ring.