9-Mercaptodethiobiotin is generated as a ligand to the [2Fe-2S]+ cluster during the reaction catalyzed by biotin synthase from Escherichia coli

J Am Chem Soc. 2012 Jun 6;134(22):9042-5. doi: 10.1021/ja3012963. Epub 2012 May 29.

Abstract

Biotin synthase catalyzes formation of the thiophane ring through stepwise substitution of a sulfur atom for hydrogen atoms at the C9 and C6 positions of dethiobiotin. Biotin synthase is a radical S-adenosylmethionine (SAM) enzyme that reductively cleaves S-adenosylmethionine, generating 5'-deoxyadenosyl radicals that initially abstract a hydrogen atom from the C9 position of dethiobiotin. We have proposed that the resulting dethiobiotinyl radical is quenched by the μ-sulfide of the nearby [2Fe-2S](2+) cluster, resulting in coupled formation of 9-mercaptodethiobiotin and a reduced [2Fe-2S](+) cluster. This reduced FeS cluster is observed by electron paramagnetic resonance spectroscopy as a mixture of two orthorhombic spin systems. In the present work, we use isotopically labeled 9-mercaptodethiobiotin and enzyme to probe the ligand environment of the [2Fe-2S](+) cluster in this reaction intermediate. Hyperfine sublevel correlation spectroscopy (HYSCORE) spectra exhibit strong cross-peaks demonstrating strong isotropic coupling of the nuclear spin with the paramagnetic center. The hyperfine coupling constants are consistent with a structural model for the reaction intermediate in which 9-mercaptodethiobiotin is covalently coordinated to the remnant [2Fe-2S](+) cluster.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biocatalysis
  • Biotin / analogs & derivatives*
  • Biotin / biosynthesis
  • Biotin / chemistry
  • Biotin / metabolism
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism*
  • Iron-Sulfur Proteins / chemistry
  • Iron-Sulfur Proteins / metabolism*
  • Ligands
  • Molecular Structure
  • Sulfurtransferases / chemistry
  • Sulfurtransferases / metabolism*

Substances

  • Escherichia coli Proteins
  • Iron-Sulfur Proteins
  • Ligands
  • 9-mercaptodethiobiotin
  • Biotin
  • Sulfurtransferases
  • biotin synthase, E coli