Ether cross-link formation in the R2-like ligand-binding oxidase

J Biol Inorg Chem. 2018 Aug;23(6):879-886. doi: 10.1007/s00775-018-1583-3. Epub 2018 Jun 26.


R2-like ligand-binding oxidases contain a dinuclear metal cofactor which can consist either of two iron ions or one manganese and one iron ion, but the heterodinuclear Mn/Fe cofactor is the preferred assembly in the presence of MnII and FeII in vitro. We have previously shown that both types of cofactor are capable of catalyzing formation of a tyrosine-valine ether cross-link in the protein scaffold. Here we demonstrate that Mn/Fe centers catalyze cross-link formation more efficiently than Fe/Fe centers, indicating that the heterodinuclear cofactor is the biologically relevant one. We further explore the chemical potential of the Mn/Fe cofactor by introducing mutations at the cross-linking valine residue. We find that cross-link formation is possible also to the tertiary beta-carbon in an isoleucine, but not to the secondary beta-carbon or tertiary gamma-carbon in a leucine, nor to the primary beta-carbon of an alanine. These results illustrate that the reactivity of the cofactor is highly specific and directed.

Keywords: Di-metal carboxylate protein; Ferritin; R2-like ligand-binding oxidase; Ribonucleotide reductase; X-ray crystallography.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Amino Acids / chemistry
  • Amino Acids / metabolism
  • Carbon / metabolism
  • Catalysis
  • Crystallization
  • Iron / metabolism
  • Ligands
  • Manganese / metabolism
  • Mass Spectrometry
  • Mutagenesis, Site-Directed
  • Oxidoreductases / chemistry
  • Oxidoreductases / genetics
  • Oxidoreductases / metabolism*


  • Amino Acids
  • Ligands
  • Manganese
  • Carbon
  • Iron
  • Oxidoreductases