Dataset Information

Peptidoglycan metabolite photoaffinity reporters reveal direct binding to intracellular pattern recognition receptors and Arf GTPases

ABSTRACT: Extensive structure-activity studies of iE-DAP and MDP have demonstrated their selective activation of NOD1- and NOD2-expressing cells, respectively. Nonetheless, the direct binding of iE-DAP and MDP to NOD1 and NOD2, respectively, as well as other proteins in mammalian cells has not been systematically investigated. To address the direct interaction of iE-DAP and MDP with NOD1 and NOD2 in living cells, we synthesized a series of peptidoglycan metabolite photoaffinity chemical reporters containing a diazirine for photocrosslinking in cells and an alkyne tag for bioorthogonal detection of covalently-labeled proteins. The analysis of these peptidoglycan metabolite photoaffinity reporters demonstrate that iE-DAP and MDP can directly bind to NOD1 and NOD2 in mammalian cells. The crosslinking of these peptidoglycan metabolite photoaffinity reporters only occur with active variants of NOD1 and NOD2. Beyond direct binding to NOD2, the active MDP photoaffinity reporter selectively crosslinked Arf GTPases in NOD2-expressing cells and did so most prominently with GTP-bound Arf6, revealing the Arf proteins as novel direct pattern recognition receptors. Additional labeling and co-precipitation experiments suggest that MDP induces a complex between NOD2 and GTP-Arf6. This study highlights the utility of peptidoglycan metabolite photoaffinity reporters for characterizing their direct binding to intracellular pattern recognition receptors and identifying unpredicted cellular cofactors. The applications of these microbial-specific metabolite reporters should enable the discovery of other receptors, transporters and regulatory enzymes in host cells and microbes.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Inflammation

SUBMITTER: Yen-Chih Wang

LAB HEAD: Howard Hang

PROVIDER: PXD012350 | Pride | 2020-05-26

REPOSITORIES: pride

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			Action	DRS
	MS184232QEP_YenChih_Hang_1-1H.raw	Raw
	MS184232QEP_YenChih_Hang_1-1L.raw	Raw
	MS184232QEP_YenChih_Hang_1-2H.raw	Raw
	MS184232QEP_YenChih_Hang_1-2L.raw	Raw
	MS184232QEP_YenChih_Hang_1-3H.raw	Raw

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Peptidoglycan Metabolite Photoaffinity Reporters Reveal Direct Binding to Intracellular Pattern Recognition Receptors and Arf GTPases.

Wang Yen-Chih YC Westcott Nathan P NP Griffin Matthew E ME Hang Howard C HC

ACS chemical biology 20190220 3

The peptidoglycan fragments γ-d-glutamyl- meso-diaminopimelic acid (iE-DAP) and muramyl-dipeptide (MDP) are microbial-specific metabolites that activate intracellular pattern recognition receptors and stimulate immune signaling pathways. While extensive structure-activity studies have demonstrated that these bacterial cell wall metabolites trigger NOD1- and NOD2-dependent signaling, their direct binding to these innate immune receptors or other proteins in mammalian cells has not been establishe ...[more]

PMID: 30735346

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