9 research outputs found
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AIG1 and ADTRP are Atypical Integral Membrane Hydrolases that Degrade Bioactive FAHFAs
Enzyme classes may contain outlier members that share mechanistic, but not sequence or structural relatedness with more common representatives. The functional annotation of such exceptional proteins can be challenging. Here, we use activity-based profiling to discover that the poorly characterized multipass transmembrane proteins AIG1 and ADTRP are atypical hydrolytic enzymes that depend on conserved threonine and histidine residues for catalysis. Both AIG1 and ADTRP hydrolyze bioactive fatty-acid esters of hydroxy-fatty acids (FAHFAs), but not other major classes of lipids. We discover multiple cell-active, covalent inhibitors of AIG1 and show that these agents block FAHFA hydrolysis in mammalian cells. These results indicate that AIG1 and ADTRP are founding members of an evolutionarily conserved class of transmembrane threonine hydrolases involved in bioactive lipid metabolism. More generally, our findings demonstrate how chemical proteomics can excavate potential cases of convergent/parallel protein evolution that defy conventional sequence- and structure-based predictions
Arylfluorosulfates Inactivate Intracellular Lipid Binding Protein(s) through Chemoselective SuFEx Reaction with a Binding Site Tyr Residue
Arylfluorosulfates
have appeared only rarely in the literature
and have not been explored as probes for covalent conjugation to proteins,
possibly because they were assumed to possess high reactivity, as
with other sulfurÂ(VI) halides. However, we find that arylfluorosulfates
become reactive only under certain circumstances, e.g., when fluoride
displacement by a nucleophile is facilitated. Herein, we explore the
reactivity of structurally simple arylfluorosulfates toward the proteome
of human cells. We demonstrate that the protein reactivity of arylfluorosulfates
is lower than that of the corresponding aryl sulfonyl fluorides, which
are better characterized with regard to proteome reactivity. We discovered
that simple hydrophobic arylfluorosulfates selectively react with
a few members of the intracellular lipid binding protein (iLBP) family.
A central function of iLBPs is to deliver small-molecule ligands to
nuclear hormone receptors. Arylfluorosulfate probe <b>1</b> reacts
with a conserved tyrosine residue in the ligand-binding site of a
subset of iLBPs. Arylfluorosulfate probes <b>3</b> and <b>4</b>, featuring a biphenyl core, very selectively and efficiently
modify cellular retinoic acid binding protein 2 (CRABP2), both in
vitro and in living cells. The X-ray crystal structure of the CRABP2–<b>4</b> conjugate, when considered together with binding site mutagenesis
experiments, provides insight into how CRABP2 might activate arylfluorosulfates
toward site-specific reaction. Treatment of breast cancer cells with
probe <b>4</b> attenuates nuclear hormone receptor activity
mediated by retinoic acid, an endogenous client lipid of CRABP2. Our
findings demonstrate that arylfluorosulfates can selectively target
single iLBPs, making them useful for understanding iLBP function
Arylfluorosulfates Inactivate Intracellular Lipid Binding Protein(s) through Chemoselective SuFEx Reaction with a Binding Site Tyr Residue
Arylfluorosulfates
have appeared only rarely in the literature
and have not been explored as probes for covalent conjugation to proteins,
possibly because they were assumed to possess high reactivity, as
with other sulfurÂ(VI) halides. However, we find that arylfluorosulfates
become reactive only under certain circumstances, e.g., when fluoride
displacement by a nucleophile is facilitated. Herein, we explore the
reactivity of structurally simple arylfluorosulfates toward the proteome
of human cells. We demonstrate that the protein reactivity of arylfluorosulfates
is lower than that of the corresponding aryl sulfonyl fluorides, which
are better characterized with regard to proteome reactivity. We discovered
that simple hydrophobic arylfluorosulfates selectively react with
a few members of the intracellular lipid binding protein (iLBP) family.
A central function of iLBPs is to deliver small-molecule ligands to
nuclear hormone receptors. Arylfluorosulfate probe <b>1</b> reacts
with a conserved tyrosine residue in the ligand-binding site of a
subset of iLBPs. Arylfluorosulfate probes <b>3</b> and <b>4</b>, featuring a biphenyl core, very selectively and efficiently
modify cellular retinoic acid binding protein 2 (CRABP2), both in
vitro and in living cells. The X-ray crystal structure of the CRABP2–<b>4</b> conjugate, when considered together with binding site mutagenesis
experiments, provides insight into how CRABP2 might activate arylfluorosulfates
toward site-specific reaction. Treatment of breast cancer cells with
probe <b>4</b> attenuates nuclear hormone receptor activity
mediated by retinoic acid, an endogenous client lipid of CRABP2. Our
findings demonstrate that arylfluorosulfates can selectively target
single iLBPs, making them useful for understanding iLBP function
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An LXR-Cholesterol Axis Creates a Metabolic Co-Dependency for Brain Cancers
Small-molecule inhibitors targeting growth factor receptors have failed to show efficacy for brain cancers, potentially due to their inability to achieve sufficient drug levels in the CNS. Targeting non-oncogene tumor co-dependencies provides an alternative approach, particularly if drugs with high brain penetration can be identified. Here we demonstrate that the highly lethal brain cancer glioblastoma (GBM) is remarkably dependent on cholesterol for survival, rendering these tumors sensitive to Liver X receptor (LXR) agonist-dependent cell death. We show that LXR-623, a clinically viable, highly brain-penetrant LXRα-partial/LXRβ-full agonist selectively kills GBM cells in an LXRβ- and cholesterol-dependent fashion, causing tumor regression and prolonged survival in mouse models. Thus, a metabolic co-dependency provides a pharmacological means to kill growth factor-activated cancers in the CNS