Clues to the mechanism of cholesterol transfer from the structure of NPC1 middle lumenal domain bound to NPC2

Li, Xiaochun; Saha, Papita; Li, Jian; Blobel, Günter; Pfeffer, Suzanne R

doi:10.1073/pnas.1611956113

Cited by 161 publications

(195 citation statements)

References 32 publications

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“…In our previous study, we proposed an NTD-SSD insertion mechanism to explain how cholesterol is transferred from the lysosomal lumen to the membrane domain (21,23). In light of the structure, we suspect that the Ω loop-Ψ loop interaction might modulate the NTD-SSD insertion to facilitate cholesterol transport.…”

Section: Resultsmentioning

confidence: 99%

“…Alternatively, or in addition, the regulation of the NTD could be affected by Lamp (lysosomalassociated membrane protein) proteins (29). After cholesterol is slipped from NPC2 to the NPC1-NTD, NPC2 changes its conformation to disassociate from the MLD-binding site (23), and the Ω loop-Ψ loop interaction might be weakened to allow the NTD to reach across the glycocalyx for delivering cholesterol to the SSD pocket in the membrane. How the NTD delivers cholesterol to the SSD in the glycocalyx is still a mystery.…”

Section: Discussionmentioning

confidence: 99%

“…Email: xli05@rockefeller.edu or blobel@ rockefeller.edu. structure of NPC1 at 4.4 Å presents the overall architecture of the full-length molecule (22); and (v) a complex of NPC1-MLD and NPC2 suggests a molecular mechanism of cholesterol transfer from NPC2 to the NTD of NPC1 (23). However, due to low resolution, our understanding of the structural details and function of the NPC1-CTD remain rudimentary.…”

Section: Significancementioning

confidence: 99%

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3.3 Å structure of Niemann–Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport

Trinh

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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103

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Niemann-Pick C1 (NPC1) and NPC2 proteins are indispensable for the export of LDL-derived cholesterol from late endosomes. Mutations in these proteins result in Niemann-Pick type C disease, a lysosomal storage disease. Despite recent reports of the NPC1 structure depicting its overall architecture, the function of its C-terminal luminal domain (CTD) remains poorly understood even though 45% of NPC disease-causing mutations are in this domain. Here, we report a crystal structure at 3.3 Å resolution of NPC1* (residues 314-1,278), which-in contrast to previous lower resolution structures-features the entire CTD well resolved. Notably, all eight cysteines of the CTD form four disulfide bonds, one of which (C909-C914) enforces a specific loop that in turn mediates an interaction with a loop of the N-terminal domain (NTD). Importantly, this loop and its interaction with the NTD were not observed in any previous structures due to the lower resolution. Our mutagenesis experiments highlight the physiological relevance of the CTD-NTD interaction, which might function to keep the NTD in the proper orientation for receiving cholesterol from NPC2. Additionally, this structure allows us to more precisely map all of the disease-causing mutations, allowing future molecular insights into the pathogenesis of NPC disease.cholesterol transport | Niemann-Pick type C disease | cysteine-rich domain | sterol-sensing domain | crystal structure

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

See 1 more Smart Citation

3.3 Å structure of Niemann–Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport

Trinh

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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103

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“…The mechanism for NPC1 function is beginning to be unraveled through biochemical and structural studies (6)(7)(8)(9)(10)(11)(12). NPC1 has 13 transmembrane helices and three large domains that project into the lumen (13).…”

mentioning

confidence: 99%

“…NPC1 has 13 transmembrane helices and three large domains that project into the lumen (13). Cholesterol-carrying NPC2 binds to the middle luminal domain (MLD), which positions the protein so that it can transfer its cholesterol to the N-terminal domain (NTD) (11). This transfer is facilitated by the formation of a channel between NPC2 and the NTD that allows cholesterol to slide from NPC2 to the NTD of NPC1 in a process known as a "hydrophobic handoff" (7).…”

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confidence: 99%

Triazoles inhibit cholesterol export from lysosomes by binding to NPC1

Trinh

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Niemann-Pick C1 (NPC1), a membrane protein of lysosomes, is required for the export of cholesterol derived from receptor-mediated endocytosis of LDL. Lysosomal cholesterol export is reportedly inhibited by itraconazole, a triazole that is used as an antifungal drug [Xu et al. (2010) Proc Natl Acad Sci USA 107:4764-4769]. Here we show that posaconazole, another triazole, also blocks cholesterol export from lysosomes. We prepared P-X, a photoactivatable cross-linking derivative of posaconazole. P-X cross-linked to NPC1 when added to intact cells. Cross-linking was inhibited by itraconazole but not by ketoconazole, an imidazole that does not block cholesterol export. Cross-linking of P-X was also blocked by U18666A, a compound that has been shown to bind to NPC1 and inhibit cholesterol export. P-X also crosslinked to purified NPC1 that was incorporated into lipid bilayer nanodiscs. In this in vitro system, cross-linking of P-X was inhibited by itraconazole, but not by U18666A. P-X cross-linking was not prevented by deletion of the N-terminal domain of NPC1, which contains the initial binding site for cholesterol. In contrast, P-X cross-linking was reduced when NPC1 contained a point mutation (P691S) in its putative sterolsensing domain. We hypothesize that the sterol-sensing domain has a binding site that can accommodate structurally different ligands.Niemann-Pick C disease | cholesterol transport | sterol-sensing domain | lipid nanodiscs | photoactivatable cross-linking

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Sterol uptake by the NPC system in eukaryotes: a Saccharomyces cerevisiae perspective

et al. 2021

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Sterols are an essential component of membranes in all eukaryotic cells and the precursor of multiple indispensable cellular metabolites. After endocytotic uptake, sterols are integrated into the lysosomal membrane by the Niemann-Pick type C (NPC) system before redistribution to other membranes. The process is driven by two proteins that, together, compose the NPC system: the lysosomal sterol shuttle protein NPC2 and the membrane protein NPC1 (named NCR1 in fungi), which integrates sterols into the lysosomal membrane. The Saccharomyces cerevisiae NPC system provides a compelling model to study the molecular mechanism of sterol integration into membranes and sterol homeostasis. This review summarizes recent advances in the field, and by interpreting available structural data, we propose a unifying conceptual model for sterol loading, transfer and transport by NPC proteins.

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Clues to the mechanism of cholesterol transfer from the structure of NPC1 middle lumenal domain bound to NPC2

Cited by 161 publications

References 32 publications

3.3 Å structure of Niemann–Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport

3.3 Å structure of Niemann–Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport

Triazoles inhibit cholesterol export from lysosomes by binding to NPC1

Sterol uptake by the NPC system in eukaryotes: a Saccharomyces cerevisiae perspective

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