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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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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
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author Saha, Piyali
Shumate, Justin L
Caldwell, Jenna G
Elghobashi-Meinhardt, Nadia
Lu, Albert
Zhang, Lichao
Olsson, Niclas E
Elias, Joshua E
Pfeffer, Suzanne R
author_facet Saha, Piyali
Shumate, Justin L
Caldwell, Jenna G
Elghobashi-Meinhardt, Nadia
Lu, Albert
Zhang, Lichao
Olsson, Niclas E
Elias, Joshua E
Pfeffer, Suzanne R
author_sort Saha, Piyali
collection PubMed
description 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.
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spelling pubmed-72287652020-05-18 Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins Saha, Piyali Shumate, Justin L Caldwell, Jenna G Elghobashi-Meinhardt, Nadia Lu, Albert Zhang, Lichao Olsson, Niclas E Elias, Joshua E Pfeffer, Suzanne R eLife Biochemistry and Chemical Biology 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. eLife Sciences Publications, Ltd 2020-05-15 /pmc/articles/PMC7228765/ /pubmed/32410728 http://dx.doi.org/10.7554/eLife.57089 Text en © 2020, Saha et al http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Biochemistry and Chemical Biology
Saha, Piyali
Shumate, Justin L
Caldwell, Jenna G
Elghobashi-Meinhardt, Nadia
Lu, Albert
Zhang, Lichao
Olsson, Niclas E
Elias, Joshua E
Pfeffer, Suzanne R
Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins
title Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins
title_full Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins
title_fullStr Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins
title_full_unstemmed Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins
title_short Inter-domain dynamics drive cholesterol transport by NPC1 and NPC1L1 proteins
title_sort inter-domain dynamics drive cholesterol transport by npc1 and npc1l1 proteins
topic Biochemistry and Chemical Biology
url 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
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