VTVH-MCD and DFT Studies of Thiolate Bonding to {FeNO}7/{FeO2}8 Complexes of Isopenicillin N Synthase: Substrate
Determination of Oxidase versus Oxygenase Activity in
Nonheme Fe Enzymes
posted on 2007-06-13, 00:00authored byChristina D. Brown, Michael L. Neidig, Matthew B. Neibergall, John D. Lipscomb, Edward I. Solomon
Isopenicillin N synthase (IPNS) is a unique mononuclear nonheme Fe enzyme that catalyzes
the four-electron oxidative double ring closure of its substrate ACV. A combination of spectroscopic
techniques including EPR, absorbance, circular dichroism (CD), magnetic CD, and variable-temperature,
variable-field MCD (VTVH-MCD) were used to evaluate the geometric and electronic structure of the
{FeNO}7 complex of IPNS coordinated with the ACV thiolate ligand. Density Function Theory (DFT)
calculations correlated to the spectroscopic data were used to generate an experimentally calibrated bonding
description of the Fe−IPNS−ACV−NO complex. New spectroscopic features introduced by the binding of
the ACV thiolate at 13 100 and 19 800 cm-1 are assigned as the NO π*(ip) → Fe dx2-y2 and S π→ Fe dx2-y2
charge transfer (CT) transitions, respectively. Configuration interaction mixes S CT character into the NO
π*(ip) → Fe dx2-y2 CT transition, which is observed experimentally from the VTVH-MCD data from this
transition. Calculations on the hypothetical {FeO2}8 complex of Fe−IPNS−ACV reveal that the configuration
interaction present in the {FeNO}7 complex results in an unoccupied frontier molecular orbital (FMO) with
correct orientation and distal O character for H-atom abstraction from the ACV substrate. The energetics
of NO/O2 binding to Fe−IPNS−ACV were evaluated and demonstrate that charge donation from the ACV
thiolate ligand renders the formation of the FeIII-superoxide complex energetically favorable, driving the
reaction at the Fe center. This single center reaction allows IPNS to avoid the O2 bridged binding generally
invoked in other nonheme Fe enzymes that leads to oxygen insertion (i.e., oxygenase function) and
determines the oxidase activity of IPNS.