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Author Notes:

Corresponding authors. J.B.B.: e-mail, jbroderick@chemistry.montana. edu; phone, (406) 994-6160; fax, (406) 994-5407. B.H.H.: vhuynh@emory.edu; phone, (404) 727-4295; fax, (404) 727-0873.

The authors thank Kaitlin Duschene for assistance in preparation of the Table of Contents illustration.


Research Funding:

This work was supported by grants from the National Institutes of Health (GM54608 to J.B.B. and GM47295 to B.H.H.)


  • Science & Technology
  • Life Sciences & Biomedicine
  • Biochemistry & Molecular Biology
  • 4FE-4S
  • GENE
  • SITE

The Iron-Sulfur Cluster of Pyruvate Formate-Lyase Activating Enzyme in Whole Cells: Cluster Interconversion and a Valence-Localized [4Fe-4S](2+) State


Journal Title:



Volume 48, Number 39


, Pages 9234-9241

Type of Work:

Article | Post-print: After Peer Review


Pyruvate formate-lyase activating enzyme (PFL-AE) catalyzes the generation of a catalytically essential glycyl radical on pyruvate formate-lyase (PFL). Purified PFL-AE contains an oxygen-sensitive, labile [4Fe-4S] cluster that undergoes cluster interconversions in vitro, with only the [4Fe-4S]+ cluster state being catalytically active. Such cluster interconversions could play a role in regulating the activity of PFL-AE, and thus of PFL, in response to oxygen levels in vivo. Here we report a Mössbauer investigation on whole cells overexpressing PFL-AE following incubation under aerobic and/or anaerobic conditions and provide evidence that PFL-AE undergoes cluster interconversions in vivo. After 2 h aerobic induction of PFL-AE expression, approximately 44% of the total iron is present in [4Fe-4S]2+ clusters, 6% in [2Fe-2S] 2+ clusters, and the remainder as noncluster FeIII (29%) and FeII (21%) species. Subsequent anaerobic incubation of the culture results in approximately 75% of the total iron being present as [4Fe-4S]2+ clusters, with no detectable [2Fe-2S]2+. Ensuing aerobic incubation of the culture converts the iron species nearly back to the original composition (42% [4Fe-4S]2+, 10% [2Fe-2S] 2+, 19% FeIII, and 29% FeII). The results provide evidence for changes in cluster composition of PFL-AE in response to the redox state of the cell. Furthermore, the Mössbauer spectra reveal that the [4Fe-4S]2+ cluster of PFL-AE in whole cells contains a valence-localized FeIIIFeII pair which has not been previously observed in the purified enzyme. Addition of certain small molecules containing adenosyl moieties, including 5′-deoxyadenosine, AMP, ADP, and methylthioadenosine, to purified PFL-AE reproduces the valence-localized state of the [4Fe-4S]2+ cluster. It is speculated that the [4Fe-4S] 2+ cluster of PFL-AE in whole cells may be coordinated by a small molecule, probably AMP, and that such coordination may protect this labile cluster from oxidative damage.

Copyright information:

© 2009 American Chemical Society.

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