RESUMEN
A branched flexible linker that incorporates a fluorescent dansyl moiety was synthesized and used to connect two high affinity NDP-alpha-MSH ligands or two low affinity MSH(4) ligands. The linker was incorporated into the conjugate by solid-phase synthesis. In vitro biological evaluations showed that potency of binding to the human melanocortin 4 receptor was not diminished for linker-ligand combinations relative to the corresponding ligand alone.
Asunto(s)
Péptidos/síntesis química , alfa-MSH/química , Cromatografía Líquida de Alta Presión , Diseño de Fármacos , Colorantes Fluorescentes , Indicadores y Reactivos , Ligandos , Espectrometría de Masas , Polietilenglicoles , Reproducibilidad de los Resultados , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
Rigid linkers of variable length were used to connect two high-affinity Nle4-D-Phe7-alpha-melanocyte stimulating hormone (NDP-alpha-MSH) or two low-affinity MSH(4) ligands. The linked peptides were synthesized by solid-phase methods. Control experiments indicate there is little or no effect of these linkers on NDP-alpha-MSH or MSH(4) binding to the human melanocortin 4 receptor (hMC4R). Tethering two high-affinity ligands gave no binding enhancement, while tethering two low-affinity ligands resulted in binding enhancement that decreased with increased linker length. Furthermore, for the low-affinity ligands, the enhancement of affinity is inversely proportional to the estimated molecular moments of inertia. These results are consistent with a model wherein binding is enhanced when the rate of ligand reattachment to the receptor is fast relative to the rate of ligand diffusion.
Asunto(s)
Reactivos de Enlaces Cruzados/química , Péptidos/química , Péptidos/metabolismo , Línea Celular , Dimerización , Humanos , Ligandos , Estructura Molecular , Unión Proteica , Receptor de Melanocortina Tipo 4/metabolismoRESUMEN
An enkephalin analogue coupled to 'aminofentanyl' has been synthesized and tested for biological activities at the mu and delta opioid receptors. Aminofentanyl which represents a structural derivative of fentanyl has been synthesized by acylation of 1-(2-phenethyl)-4-(N-anilino)piperidine with phthaloyl protected beta-alaninyl chloride in the presence of DIPEA, followed by deprotection with hydrazine hydrate. Aminofentanyl has also been successfully acylated with ethyl isocyanate, various acid anhydrides, to further investigate structure-activity relationships of these new fentanyl derivatives. Among the new derivatives compound 7 which carries a Tyr-D-Ala-Gly-Phe opioid message sequence showed good opioid affinity (1 nM at both delta and mu opioid receptors) and bioactivity (34.9 nM in MVD and 42 nM in GPI/LMMP bioassays).