A Global Map of Lipid-Binding Proteins and Their Ligandability in Cells

Abstract

Summary Lipids play central roles in physiology and disease, where their structural, metabolic, and signaling functions often arise from interactions with proteins. Here, we describe a set of lipid-based chemical proteomic probes and their global interaction map in mammalian cells. These interactions involve hundreds of proteins from diverse functional classes and frequently occur at sites of drug action. We determine the target profiles for several drugs across the lipidinteraction proteome, revealing that its ligandable content extends far beyond traditionally defined categories of druggable proteins. In further support of this finding, we describe a selective ligand for the lipid-binding protein nucleobindin-1 (NUCB1) and show that this compound perturbs the hydrolytic and oxidative metabolism of endocannabinoids in cells. The described chemical proteomic platform thus provides an integrated path to both discover and pharmacologically characterize a wide range of proteins that participate in lipid pathways in cells.

Trifunctional lipid probes for comprehensive studies of single lipid species in living cells

“…3A. As noted in previous studies using bifunctional lipids (20,21), a high overlap in the interactome of different lipids was observed; this may be attributed to unspecific cross-linking of highly abundant proteins with either probe or background binding to the beads during the pull-down procedure, highlighting the need for control lipids. By grouping the resulting proteins according to the ratio of their peptide spectral matches over control lipids, we were able to identify two subsets of proteins, which interacted preferentially with either TFS (Fig.…”

“…Here, we benefited from the major advantage of trifunctional lipids to circumvent lipid metabolism and identify scarcer or very short-lived lipid−protein interactions. In the absence of previous proteomic screens with DAG as bait, we compared proteins identified using TFDAG to hits from previous screens performed with bifunctional endocannabinoid probes (21). These probes contain arachidonic acid just as TFDAG does.…”

“…Biotinylation of cross-linked lipid−protein conjugates enables their enrichment and identification of lipidinteracting proteins. To date, bifunctional lipids are one of the few methods to screen for lipid−protein interactions in living cells (18)(19)(20)(21). Alternatively, bifunctional lipids can be used to visualize lipid localization by click reaction with a fluorophore (1,18,20).…”

A search for ceramide binding proteins using bifunctional lipid analogs yields CERT-related protein StarD7

“…Moreover, interactions where the lipid has to enter a deep hydrophobic binding pocket within the protein are likely to be missed. In recent years, bifunctional lipid analogs have emerged as promising new tools to circumvent some of these disadvantages, by guest, on May 11, 2018 www.jlr.org Downloaded from enabling a global profiling of lipid-protein interactions in living cells (25)(26)(27)(28). Bifunctional lipids possess a small diazirine group to allow photo-crosslinking with their protein interaction partners and a terminal alkyne or clickable group for functionalization.…”

Mice lacking have reduced signs of metabolic syndrome and non-alcoholic fatty liver disease

“…The same authors observed that early loss of Arv1 caused increases in plasma PL, cholesterol, and LDL-C (low density apolipoprotein-cholesterol) levels, and a decrease in HDL-C (high density apolipoprotein-cholesterol) levels (20). Interestingly, human ARV1 possesses in vitro lipid binding activity, as it binds sterol intermediates (21), several fatty acid species (22), and specific phospholipids and cholesterol (H-Y…”