Loss of Iron-Sulfur Clusters From Biotin Synthase as a Result of Catalysis Promotes Unfolding and Degradation

Arch Biochem Biophys. 2008 Mar 1;471(1):32-41. doi: 10.1016/j.abb.2007.12.001. Epub 2007 Dec 10.


Biotin synthase (BioB) is an S-adenosylmethionine radical enzyme that catalyzes addition of sulfur to dethiobiotin to form the biotin thiophane ring. In vitro, Escherichia coli BioB is active for only one turnover, during which the [2Fe-2S]2+ cluster is destroyed, one sulfide from the cluster is incorporated as the biotin thiophane sulfur, while Fe2+ ions and the remaining S2- ion are released from the protein. The present work examines the fate of the protein following the loss of the FeS clusters. We examine the quaternary structure and thermal stability of active and inactive states of BioB, and find that loss of either the [4Fe-4S]2+ or [2Fe-2S]2+ clusters results in destabilization but not global unfolding of BioB. Using susceptibility to limited proteolysis as a guide, we find that specific regions of the protein appear to be transiently unfolded following loss of these clusters. We also examine the in vivo degradation of biotin synthase during growth in low-iron minimal media and find that BioB is degraded by an apparent ATP-dependent proteolysis mechanism that sequentially cleaves small fragments starting at the C-terminus. BioB appears to be resistant to degradation and capable of multiple turnovers only under high-iron conditions that favor repair of the FeS clusters, a process most likely mediated by the Isc or Suf iron-sulfur cluster assembly systems.

MeSH terms

  • Apoenzymes / antagonists & inhibitors
  • Apoenzymes / chemistry
  • Apoenzymes / metabolism
  • Catalysis
  • Culture Media / metabolism
  • Dimerization
  • Enzyme Stability
  • Escherichia coli Proteins / antagonists & inhibitors
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / metabolism*
  • Gene Deletion
  • Gene Targeting
  • Iron-Sulfur Proteins / antagonists & inhibitors
  • Iron-Sulfur Proteins / chemistry*
  • Iron-Sulfur Proteins / metabolism*
  • Protein Denaturation
  • Protein Folding*
  • Protein Structure, Quaternary
  • Sulfurtransferases / antagonists & inhibitors
  • Sulfurtransferases / chemistry*
  • Sulfurtransferases / metabolism*
  • Thermodynamics


  • Apoenzymes
  • Culture Media
  • Escherichia coli Proteins
  • Iron-Sulfur Proteins
  • Sulfurtransferases
  • biotin synthetase