Chemoproteomic profiling reveals cellular targets of nitro-fatty acids

Abstract

Nitro-fatty acids are a class of endogenous electrophilic lipid mediators with anti-inflammatory and cytoprotective effects in a wide range of inflammatory and fibrotic disease models. While these beneficial biological effects of nitro-fatty acids are mainly attributed to their ability to form covalent adducts with proteins, only a small number of proteins are known to be nitro-alkylated and the scope of protein nitro-alkylation remains undetermined. Here we describe the synthesis and application of a clickable nitro-fatty acid probe for the detection and first global identification of mammalian proteins that are susceptible to nitro-alkylation. 184 high confidence nitro-alkylated proteins were identified in THP1 macrophages, majority of which are novel targets of nitro-fatty acids, including extended synaptotagmin 2 (ESYT2), signal transducer and activator of transcription 3 (STAT3), toll-like receptor 2 (TLR2), retinoid X receptor alpha (RXRα) and glucocorticoid receptor (NR3C1). In particular, we showed that 9-nitro-oleate covalently modified and inhibited dexamethasone binding to NR3C1. Bioinformatic analyses revealed that nitro-alkylated proteins are highly enriched in endoplasmic reticulum and transmembrane proteins, and are overrepresented in lipid metabolism and transport pathways. This study significantly expands the scope of protein substrates targeted by nitro-fatty acids in living cells and provides a useful resource towards understanding the pleiotropic biological roles of nitro-fatty acids as signaling molecules or as multi-target therapeutic agents.

Nitro-Oleic Acid-Mediated Nitroalkylation Modulates the Antioxidant Function of Cytosolic Peroxiredoxin Tsa1 during Heat Stress in Saccharomyces cerevisiae

“…The complexity involved in detecting a PTM as nitroalkylation must be highlighted. To date, this modification had only been identified and characterized in in vitro studies [61]. This makes the present research work the first study to detect the in vivo NO 2 -OA-mediated nitroalkylation.…”

Nitro Fatty Acids (NO2-FAs): An Emerging Class of Bioactive Fatty Acids

“…In 2021, Zhang et al reported the synthesis of 9-nitroeicos-9-en-19-ynoic acid (99), a derivative of 9-NO 2 -OA, where a terminal triple bond exists (Scheme 25) [119]. Compound 99 may serve as a tool of detecting in general which mammalian proteins can form covalent adducts with the electrophilic center of this NO 2 -FA.…”

Multi-omics Investigation into the Mechanism of Action of an Anti-tubercular Fatty Acid Analogue

“…Moreover, the clickable nature of DA-CB may be leveraged to trace the cellular fate of the modified fatty acid or any derived synthetic intermediate. 20…”

“…In addition to its efficacy, DA-CB is an intriguing lead compound because a strained cyclic alkene is present. This feature enables selective addressing of DA-CB or any derived biosynthetic intermediate via a “click” reaction, − providing a handle with which to decipher the location of the fatty acid analogue or derived conjugates within the mycobacterial cell.…”

“…Our results suggest that DA-CB and related molecules have the potential for development as TB therapeutics. Moreover, the clickable nature of DA-CB may be leveraged to trace the cellular fate of the modified fatty acid or any derived synthetic intermediate …”