RESUMEN
Huntington's disease is a neurodegenerative disorder associated with the expansion of the polyglutamine tract in the exon-1 domain of the huntingtin protein (htte1). Above a threshold of 37 glutamine residues, htte1 starts to aggregate in a nucleation-dependent manner. A 17-residue N-terminal fragment of htte1 (N17) has been suggested to play a crucial role in modulating the aggregation propensity and toxicity of htte1. Here we identify N17 as a potential target for novel therapeutic intervention using the molecular tweezer CLR01. A combination of biochemical experiments and computer simulations shows that binding of CLR01 induces structural rearrangements within the htte1 monomer and inhibits htte1 aggregation, underpinning the key role of N17 in modulating htte1 toxicity.
Asunto(s)
Hidrocarburos Aromáticos con Puentes/farmacología , Proteína Huntingtina/antagonistas & inhibidores , Organofosfatos/farmacología , Hidrocarburos Aromáticos con Puentes/química , Exones , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Simulación de Dinámica Molecular , Estructura Molecular , Organofosfatos/química , Agregado de Proteínas/efectos de los fármacosRESUMEN
Huntington's disease is caused by a CAG trinucleotide expansion mutation in the Huntingtin gene that leads to an artificially long polyglutamine sequence in the Huntingtin protein. A key feature of the disease is the intracellular aggregation of the Huntingtin exon 1 protein (Httex1) into micrometer sized inclusion bodies. The aggregation process of Httex1 has been extensively studied in vitro, however, the crucial early events of nucleation and aggregation in the cell remain elusive. Here, we studied the conformational dynamics and self-association of Httex1 by in-cell experiments using laser-induced temperature jumps and analytical ultracentrifugation. Both short and long polyglutamine variants of Httex1 underwent an apparent temperature-induced conformational collapse. The temperature jumps generated a population of kinetically trapped species selectively for the longer polyglutamine variants of Httex1 proteins. Their occurrence correlated with the formation of inclusion bodies suggesting that such species trigger further self-association.
Asunto(s)
Proteína Huntingtina/química , Proteína Huntingtina/metabolismo , Agregación Patológica de Proteínas/fisiopatología , Proteína Huntingtina/genética , Enfermedad de Huntington/fisiopatología , Técnicas In Vitro , Cuerpos de Inclusión/metabolismo , Rayos Láser , Modelos Moleculares , Mutación , Péptidos/química , Pliegue de Proteína , Estructura Terciaria de Proteína , Temperatura , UltracentrifugaciónRESUMEN
The role of polyglutamine (polyQ) tract on protein stability and disease pathology remains ambiguous. We monitored the unfolding/refolding patterns of huntingtin proteins with varying polyQ lengths. In the presence of urea, minor differences in unfolding and refolding efficiencies were observed. However, in the presence of guanidinium hydrochloride, the protein with a longer polyQ stretch was able to regain its secondary but not tertiary structure on step-wise removal of denaturant. Thus, in case of Huntington's disease, the higher aggregation propensity of the mutant protein is likely to be due to the lower stability of the protein due to elongated polyQ tract.