Functional diversity of the rhodanese homology domain: the Escherichia coli ybbB gene encodes a selenophosphate-dependent tRNA 2-selenouridine synthase

J Biol Chem. 2004 Jan 16;279(3):1801-9. doi: 10.1074/jbc.M310442200. Epub 2003 Oct 31.


Escherichia coli has eight genes predicted to encode sulfurtransferases having the active site consensus sequence Cys-Xaa-Xaa-Gly. One of these genes, ybbB, is frequently found within bacterial operons that contain selD, the selenophosphate synthetase gene, suggesting a role in selenium metabolism. We show that ybbB is required in vivo for the specific substitution of selenium for sulfur in 2-thiouridine residues in E. coli tRNA. This modified tRNA nucleoside, 5-methylaminomethyl-2-selenouridine (mnm(5)se(2)U), is located at the wobble position of the anticodons of tRNA(Lys), tRNA(Glu), and tRNA(1)(Gln). Nucleoside analysis of tRNAs from wild-type and ybbB mutant strains revealed that production of mnm(5)se(2)U is lost in the ybbB mutant but that 5-methylaminomethyl-2-thiouridine, the mnm(5)se(2)U precursor, is unaffected by deletion of ybbB. Thus, ybbB is not required for the initial sulfurtransferase reaction but rather encodes a 2-selenouridine synthase that replaces a sulfur atom in 2-thiouridine in tRNA with selenium. Purified 2-selenouridine synthase containing a C-terminal His(6) tag exhibited spectral properties consistent with tRNA bound to the enzyme. In vitro mnm(5)se(2)U synthesis is shown to be dependent on 2-selenouridine synthase, SePO(3), and tRNA. Finally, we demonstrate that the conserved Cys(97) (but not Cys(96)) in the rhodanese sequence motif Cys(96)-Cys(97)-Xaa-Xaa-Gly is required for 2-selenouridine synthase in vivo activity. These data are consistent with the ybbB gene encoding a tRNA 2-selenouridine synthase and identifies a new role for the rhodanese homology domain in enzymes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Cloning, Molecular
  • Escherichia coli / enzymology*
  • Escherichia coli Proteins / chemistry*
  • Molecular Sequence Data
  • Phosphates / physiology*
  • Selenium / metabolism
  • Selenium Compounds
  • Sulfurtransferases / chemistry*
  • Sulfurtransferases / genetics
  • Sulfurtransferases / physiology
  • Thiosulfate Sulfurtransferase / chemistry*
  • Thiosulfate Sulfurtransferase / physiology


  • Escherichia coli Proteins
  • Phosphates
  • Selenium Compounds
  • selenophosphate
  • Sulfurtransferases
  • tRNA 2-selenouridine synthase
  • Thiosulfate Sulfurtransferase
  • Selenium