Mechanistic Studies of Niemann–Pick C1

Research Assistant Professor Feiran Lu, Ph.D.
The University of Texas Southwestern Medical Center, Dallas, Texas, USA
11 May 2017 - 3:00pm
Rountree Room 356, Level 3, Biological Sciences Building D26

This seminar will highlight recent mechanistic insights into Niemann-Pick C1 (NPC1), a lysosomal membrane protein that exports cholesterol derived from receptor-mediated uptake of Low-Density Lipoprotein. Lysosomal cholesterol export is inhibited by nanomolar concentrations of U18666A, a cationic sterol, and by itraconazole, a triazole that is used as an antifungal drug. To identify the target of U18666A and triazoles, we synthesized photocrosslinkable derivatives of these compounds by attaching a benzophenone that permits ultraviolet-induced crosslinking: U-X based on U18666A and P-X based on posaconazole, another triazole. When added to CHO cells, U-X and P-X crosslinked to NPC1. Crosslinking was blocked by chemical derivatives that block cholesterol export, but not derivatives lacking blocking activity. Crosslinking was also prevented by point mutation in the sterol-sensing domain (SSD) of NPC1, but not by point mutation or deletion of the N-terminal domain (NTD). Furthermore, P-X also cross-linked 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. These data suggest that the SSD contains a site required for cholesterol export distinct from the cholesterol-binding site in the NTD and that the SSD has a binding site that can accommodate structurally different ligands.

Biography: Feiran Lu, Ph.D. is a Research Assistant Professor at in the Department of Molecular Genetics at The University of Texas Southwestern Medical Center at Dallas, TX in the United States. Her fields of expertise include biochemistry and structural biology. During graduate school at Tsinghua University, she solved high resolution crystal structures of the membrane protein transporters AdiC, UraA, and FucP in the laboratory of Yigong Shi, Ph.D. As a postdoctoral fellow in the Brown/Goldstein Laboratory, she delved deeper into the systematic study of a membrane protein cholesterol transporter named Niemann-Pick C1 (NPC1). Dr. Lu found two sets of compounds named U18666A and triazole drugs that, via biochemical and chemical biological discoveries, were found to directly inhibit NPC1. Now, as a newly minted Research Assistant Professor of Molecular Genetics, she is conducting further investigation into the NPC1 protein.