An Untargeted Approach for Revealing Electrophilic Metabolites.

ABSTRACT: Reactive electrophilic intermediates such as coenzyme A esters play central roles in metabolism but are difficult to detect with conventional strategies. Here, we introduce hydroxylamine-based stable isotope labeling to convert reactive electrophilic intermediates into stable derivatives that are easily detectable via LC-MS. In the model system Caenorhabditis elegans, parallel treatment with ¹⁴NH₂OH and ¹⁵NH₂OH revealed >1000 labeled metabolites, e.g., derived from peptide, fatty acid, and ascaroside pheromone biosyntheses. Results from NH₂OH treatment of a pheromone biosynthesis mutant, acox-1.1, suggested upregulation of thioesterase activity, which was confirmed by gene expression analysis. The upregulated thioesterase contributes to the biosynthesis of a specific subset of ascarosides, determining the balance of dispersal and attractive signals. These results demonstrate the utility of NH₂OH labeling for investigating complex biosynthetic networks. Initial results with Aspergillus and human cell lines indicate applicability toward uncovering reactive metabolomes in diverse living systems.

SUBMITTER: Yu Y 

PROVIDER: S-EPMC7852410 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature