Bacterial flavoprotein monooxygenase YxeK salvages toxic S-(2-succino)-adducts via oxygenolytic C-S bond cleavage

FEBS J. 2022 Feb;289(3):787-807. doi: 10.1111/febs.16193. Epub 2021 Oct 4.


Thiol-containing nucleophiles such as cysteine react spontaneously with the citric acid cycle intermediate fumarate to form S-(2-succino)-adducts. In Bacillus subtilis, a salvaging pathway encoded by the yxe operon has recently been identified for the detoxification and exploitation of these compounds as sulfur sources. This route involves acetylation of S-(2-succino)cysteine to N-acetyl-2-succinocysteine, which is presumably converted to oxaloacetate and N-acetylcysteine, before a final deacetylation step affords cysteine. The critical oxidative cleavage of the C-S bond of N-acetyl-S-(2-succino)cysteine was proposed to depend on the predicted flavoprotein monooxygenase YxeK. Here, we characterize YxeK and verify its role in S-(2-succino)-adduct detoxification and sulfur metabolism. Detailed biochemical and mechanistic investigation of YxeK including 18 O-isotope-labeling experiments, homology modeling, substrate specificity tests, site-directed mutagenesis, and (pre-)steady-state kinetics provides insight into the enzyme's mechanism of action, which may involve a noncanonical flavin-N5-peroxide species for C-S bond oxygenolysis.

Keywords: flavin-N5-peroxide; flavin-dependent monooxygenase; oncometabolite; oxidoreductase; oxygenolytic bond cleavage.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylation
  • Bacillus subtilis / genetics
  • Bacillus subtilis / metabolism
  • Cysteine / analogs & derivatives*
  • Cysteine / genetics*
  • Cysteine / metabolism
  • Flavins / genetics
  • Flavins / metabolism
  • Flavoproteins / genetics*
  • Flavoproteins / metabolism
  • Fumarates / metabolism
  • Kinetics
  • Mixed Function Oxygenases / genetics*
  • Models, Chemical
  • Mutagenesis, Site-Directed
  • Operon / genetics
  • Substrate Specificity / genetics
  • Sulfhydryl Compounds / metabolism


  • Flavins
  • Flavoproteins
  • Fumarates
  • S-(2-succinyl)cysteine
  • Sulfhydryl Compounds
  • Mixed Function Oxygenases
  • Cysteine