Evaluation of a concerted vs. sequential oxygen activation mechanism in α-ketoglutarate-dependent nonheme ferrous enzymes

Serra Goudarzi, Shyam R Iyer, Jeffrey T Babicz, James J Yan, Günther H.J. Peters, Hans Erik Mølager Christensen, Britt Hedman, Keith O Hodgson, Edward I Solomon*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

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Abstract

Determining the requirements for efficient oxygen (O2) activation is key to understanding how enzymes maintain efficacy and mitigate unproductive, often detrimental reactivity. For the α-ketoglutarate (αKG)-dependent nonheme iron enzymes, both a concerted mechanism (both cofactor and substrate binding prior to reaction with O2) and a sequential mechanism (cofactor binding and reaction with O2 precede substrate binding) have been proposed. Deacetoxycephalosporin C synthase (DAOCS) is an αKG-dependent nonheme iron enzyme for which both of these mechanisms have been invoked to generate an intermediate that catalyzes oxidative ring expansion of penicillin substrates in cephalosporin biosynthesis. Spectroscopy shows that, in contrast to other αKG-dependent enzymes (which are six coordinate when only αKG is bound to the FeII), αKG binding to FeII-DAOCS results in ∼45% five-coordinate sites that selectively react with O2 relative to the remaining six-coordinate sites. However, this reaction produces an FeIII species that does not catalyze productive ring expansion. Alternatively, simultaneous αKG and substrate binding to FeII-DAOCS produces five-coordinate sites that rapidly react with O2 to form an FeIV=O intermediate that then reacts with substrate to produce cephalosporin product. These results demonstrate that the concerted mechanism is operative in DAOCS and by extension, other nonheme iron enzymes.
Original languageEnglish
JournalProceedings of the National Academy of Sciences of the United States of America
Volume117
Issue number10
Pages (from-to)5152-5159
ISSN0027-8424
DOIs
Publication statusPublished - 2020

Keywords

  • Oxygen activation
  • Concerted machanism
  • Metalloenzymes
  • Oxygenase chemistry

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