Coordination of adenosylmethionine to a unique iron site of the [4Fe-4S] of pyruvate formate-lyase activating enzyme: A Mössbauer spectroscopic study

Carsten Krebs, William E. Broderick, Timothy F. Henshaw, Joan B. Broderick, Boi Hanh Huynh

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107 Scopus citations

Abstract

Pyruvate formate-lyase activating enzyme (PFL-AE) generates the catalytically essential glycyl radical of PFL. It is a member of the so-called "radical-SAM superfamily" of enzymes that use a [4Fe-4S] cluster and S-adenosylmethionine (AdoMet or SAM) to catalyze diverse radical-mediated reactions. Evidence suggests that this class of enzymes operate by common initial steps involving the generation of an AdoMet-derived adenosyl radical intermediate, of which the mechanism remains unresolved. The three-cysteine CX3CX2C cluster-binding motif common to all members of this superfamily suggests a unique Fe site in the [4Fe-4S] cluster, which presumably interacts with AdoMet to effect the reductive cleavage and radical generation. Here we employ a dual-iron-isotope (56Fe/57Fe) approach to demonstrate the existence of a unique Fe site in the [4Fe-4S] cluster of PFL-AE by Mössbauer spectroscopy. Coordination of AdoMet to this unique Fe site was made evident by the observation of a substantial increase in the isomer shift (δ) of the Mössbauer spectrum associated with the unique Fe site: δ = 0.42 mm/s in the absence of AdoMet increases to δ = 0.72 mm/s in the presence of AdoMet. Further, the Mössbauer data show that the binding of AdoMet to the unique Fe site occurs in the [4Fe-4S]2+ state, prior to the injection of the reducing equivalent required for catalysis. This observation indicates that AdoMet coordination is a necessary prerequisite to adenosyl radical generation.

Original languageEnglish (US)
Pages (from-to)912-913
Number of pages2
JournalJournal of the American Chemical Society
Volume124
Issue number6
DOIs
StatePublished - Feb 12 2002

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

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