RESUMEN
Stimulation of adipose tissue thermogenesis is regarded as a promising avenue in the treatment of obesity. However, pharmacologic engagement of this process has proven difficult. Using the Connectivity Map (CMap) approach, we identified the phytochemical hyperforin (HPF) as an anti-obesity agent. We found that HPF efficiently promoted thermogenesis by stimulating AMPK and PGC-1α via a Ucp1-dependent pathway. Using LiP-SMap (limited proteolysis-mass spectrometry) combined with a microscale thermophoresis assay and molecular docking analysis, we confirmed dihydrolipoamide S-acetyltransferase (Dlat) as a direct molecular target of HPF. Ablation of Dlat significantly attenuated HPF-mediated adipose tissue browning both in vitro and in vivo. Furthermore, genome-wide association study analysis indicated that a variation in DLAT is significantly associated with obesity in humans. These findings suggest that HPF is a promising lead compound in the pursuit of a pharmacological approach to promote energy expenditure in the treatment of obesity.
Asunto(s)
Tejido Adiposo Pardo/metabolismo , Tejido Adiposo Blanco/metabolismo , Floroglucinol/análogos & derivados , Transducción de Señal/efectos de los fármacos , Terpenos/farmacología , Termogénesis/efectos de los fármacos , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Sitios de Unión , Frío , Acetiltransferasa de Residuos Dihidrolipoil-Lisina/química , Acetiltransferasa de Residuos Dihidrolipoil-Lisina/metabolismo , Humanos , Hypericum/química , Hypericum/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Simulación del Acoplamiento Molecular , Obesidad/tratamiento farmacológico , Obesidad/metabolismo , Obesidad/patología , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/metabolismo , Floroglucinol/química , Floroglucinol/metabolismo , Floroglucinol/farmacología , Floroglucinol/uso terapéutico , Terpenos/química , Terpenos/metabolismo , Terpenos/uso terapéutico , Termogénesis/genética , Proteína Desacopladora 1/genética , Proteína Desacopladora 1/metabolismo , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Small-angle scattering of X-rays (SAXS) is an established method for low-resolution structural characterization of biological macromolecules in solution. Being complementary to the high resolution methods (X-ray crystallography and NMR), SAXS is often used in combination with them. The technique provides overall three-dimensional structures using ab initio reconstructions and hybrid modeling, and allows one to quantitatively characterize equilibrium mixtures as well as flexible systems. Recent progress in SAXS instrumentation, most notably, high brilliance synchrotron sources, has paved the way for high throughput automated SAXS studies allowing screening of external conditions (pH, temperature, ligand binding etc.). The modern approaches for SAXS data analysis are presented in this review including rapid characterization of macromolecular solutions in terms of low-resolution shapes, validation of high-resolution models in close-to-native conditions, quaternary structure analysis of complexes and quantitative description of the oligomeric composition in mixtures. Practical aspects of SAXS as a standalone tool and its combinations with other structural, biophysical or bioinformatics methods are reviewed. The capabilities of the technique are illustrated by a selection of recent applications for the studies of biological molecules. Future perspectives on SAXS and its potential impact to structural molecular biology are discussed.