Activity-Based Protein Profiling Identifies Protein Disulfide-Isomerases As Target Proteins of the Volatile Salinilactones.
Advanced science(2024)
Abstract
The salinilactones, volatile marine natural products secreted from Salinispora arenicola, feature a unique [3.1.0]-lactone ring system and cytotoxic activities through a hitherto unknown mechanism. To find their molecular target, an activity-based protein profiling with a salinilactone-derived probe is applied that disclosed the protein disulfide-isomerases (PDIs) as the dominant mammalian targets of salinilactones, and thioredoxin (TRX1) as secondary target. The inhibition of protein disulfide-isomerase A1 (PDIA1) and TRX1 is confirmed by biochemical assays with recombinant proteins, showing that (1S,5R)-salinilactone B is more potent than its (1R,5S)-configured enantiomer. The salinilactones bound covalently to C53 and C397, the catalytically active cysteines of the isoform PDIA1 according to tandem mass spectrometry. Reactions with a model substrate demonstrated that the cyclopropyl group is opened by an attack of the thiol at C6. Fluorophore labeling experiments showed the cell permeability of a salinilactone-BODIPY (dipyrrometheneboron difluoride) conjugate and its co-localization with PDIs in the endoplasmic reticulum. The study is one of the first to pinpoint a molecular target for a volatile microbial natural product, and it demonstrates that salinilactones can achieve high selectivity despite their small size and intrinsic reactivity. Molecular targets of volatile natural products are largely unknown. The identification of protein disulfide-isomerases (PDIs) are presented as a target protein family of the salinilactones, volatile metabolites produced by marine Salinispora bacteria. The unnatural (1S,5R)-salinilactone B enantiomer exhibits high target selectivity by binding covalently to protein disulfide-isomerase A1 (PDIA1) at C53 and C397 residues and localizes at the endoplasmic reticulum of A549 cells. image
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Key words
activity-based protein profiling,natural products,protein disulfide-isomerases,salinilactones,volatiles
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