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Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins

Transport of LDL-derived cholesterol from lysosomes into the cytoplasm requires NPC1 protein; NPC1L1 mediates uptake of dietary cholesterol. We introduced single disulfide bonds into NPC1 and NPC1L1 to explore the importance of inter-domain dynamics in cholesterol transport. Using a sensitive method...

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Detalles Bibliográficos
Autores principales: Saha, Piyali, Shumate, Justin L, Caldwell, Jenna G, Elghobashi-Meinhardt, Nadia, Lu, Albert, Zhang, Lichao, Olsson, Niclas E, Elias, Joshua E, Pfeffer, Suzanne R
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7228765/
https://www.ncbi.nlm.nih.gov/pubmed/32410728
http://dx.doi.org/10.7554/eLife.57089
Descripción
Sumario:Transport of LDL-derived cholesterol from lysosomes into the cytoplasm requires NPC1 protein; NPC1L1 mediates uptake of dietary cholesterol. We introduced single disulfide bonds into NPC1 and NPC1L1 to explore the importance of inter-domain dynamics in cholesterol transport. Using a sensitive method to monitor lysosomal cholesterol efflux, we found that NPC1’s N-terminal domain need not release from the rest of the protein for efficient cholesterol export. Either introducing single disulfide bonds to constrain lumenal/extracellular domains or shortening a cytoplasmic loop abolishes transport activity by both NPC1 and NPC1L1. The widely prescribed cholesterol uptake inhibitor, ezetimibe, blocks NPC1L1; we show that residues that lie at the interface between NPC1L1's three extracellular domains comprise the drug’s binding site. These data support a model in which cholesterol passes through the cores of NPC1/NPC1L1 proteins; concerted movement of various domains is needed for transfer and ezetimibe blocks transport by binding to multiple domains simultaneously.