Magnetic circular dichroism spectroscopy as a probe of the geometric and electronic structure of non-heme ferrous enzymes

Solomon, Edward I.; Pavel, Elizabeth G.; Loeb, Kelly E.; Campochiaro, Cecelia

doi:10.1016/0010-8545(95)01150-n

Cited by 217 publications

(487 citation statements)

References 164 publications

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“…Analysis of the energies and splitting pattern of CD͞MCD bands provides insight into the geometric and electronic structures of the ferrous active sites in HmaS and HPPD (18,19). The 278 K CD spectrum of apoHmaS is featureless (Fig.…”

Section: Results and Analysismentioning

confidence: 99%

Spectroscopic and electronic structure studies of aromatic electrophilic attack and hydrogen-atom abstraction by non-heme iron enzymes

Neidig

Decker

Choroba

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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(4-Hydroxy)mandelate synthase (HmaS) and (4-hydroxyphenyl)pyruvate dioxygenase (HPPD) are two ␣-keto acid dependent mononuclear non-heme iron enzymes that use the same substrate, (4-hydroxyphenyl)pyruvate, but exhibit two different general reactivities. HmaS performs hydrogen-atom abstraction to yield benzylic hydroxylated product (S)-(4-hydroxy)mandelate, whereas HPPD utilizes an electrophilic attack mechanism that results in aromatic hydroxylated product homogentisate. These enzymes provide a unique opportunity to directly evaluate the similarities and differences in the reaction pathways used for these two reactivities. An Fe II methodology using CD, magnetic CD, and variable-temperature, variable-field magnetic CD spectroscopies was applied to HmaS and compared with that for HPPD to evaluate the factors that affect substrate interactions at the active site and to correlate these to the different reactivities exhibited by HmaS and HPPD to the same substrate. Combined with density functional theory calculations, we found that HmaS and HPPD have similar substrate-bound complexes and that the role of the protein pocket in determining the different reactivities exhibited by these enzymes (hydrogen-atom abstraction vs. aromatic electrophilic attack) is to properly orient the substrate, allowing for ligand field geometric changes along the reaction coordinate. Elongation of the Fe IV AO bond in the transition state leads to dominant Fe III OO •؊ character, which significantly contributes to the reactivity with either the aromatic -system or the COH -bond. magnetic circular dichroism ͉ density functional calculations ͉ reaction coordinates

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Section: Results and Analysismentioning

confidence: 99%

Spectroscopic and electronic structure studies of aromatic electrophilic attack and hydrogen-atom abstraction by non-heme iron enzymes

Neidig

Decker

Choroba

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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151

242

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“…11 In the previous section the terms contributing to an MCD transition have been discussed. In particular, the C-term contribution is greatly enhanced at low temperatures and high fields (see Equation (1)).…”

Section: Vtvh MCDmentioning

confidence: 99%

Magnetic Circular Dichroism of Paramagnetic Species

Solomon

Neidig

Schenk

2003

Comprehensive Coordination Chemistry II

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MCD spectroscopy combines the CD experiment with a longitudinal magnetic field, where the application of the magnetic field induces optical activity in any material so that all substances exhibit MCD activity. MCD probes the Zeeman splittings in the ground and excited states and the field-induced mixing between states. MCD intensity for a transition from a ground state |Ai to an excited state |Ji is given by Equation (1):for broadband transitions when the MCD intensity is linear with field. Here, ÁA is the fielddependent difference between left-and right-circularly polarized light (lcp and rcp, respectively) absorption, E ¼ h#, is the electric permeability, C is the concentration, l is the path length, n is 339

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“…A variable-temperature, variable-field magnetic circular dichroism (VTVH MCD) methodology we developed in other studies was applied to probe this ferrous site. [9][10][11] From figure 2A dark blue, there is a characteristic feature at 8900 cm −1 in the MCD spectrum corresponding to Fe(II) binding with a high affinity (K B > 10 5 M −1 , from MCD titration studies) to a 6 coordinate site in the protein. 12 In the light blue spectrum this feature is eliminated and a peak at 9700 cm −1 , corresponding to aqueous Fe(II) (green) is observed when Zn(II) is first bound to the substrate site, inhibiting ferroxidase activity.…”

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confidence: 99%

O2 Reduction to H2O by the multicopper oxidases

2008

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In nature the four electron reduction of O 2 to H 2 O is carried out by Cytochrome c Oxidase (CcO) and the multicopper oxidases (MCOs). In the former, Cytochrome c provides electrons for pumping protons to produce a gradient for ATP synthesis, while in the MCOs the function is the oxidation of substrates, either organic or metal ions. In the MCOs the reduction of O 2 is carried out at a trinuclear Cu cluster (TNC). Oxygen intermediates have been trapped which exhibit unique spectroscopic features that reflect novel geometric and electronic structures. These intermediates have both intact and cleaved O-O bonds, allowing the reductive cleavage of the O-O bond to be studied in detail both experimentally and computationally. These studies show that the topology of the TNC provides a unique geometric and electronic structure particularly suited to carry out this key reaction in Nature.The multicopper oxidases (MCOs) couple four 1-electron oxidations of substrate to the four electron reductive cleavage of the O-O bond of dioxygen using a minimum of four Cu atoms (table 1). 1,2 Among these four Cu's is a type 1 (T1) or blue Cu site, characterized by an intense S Cys → Cu(II) charge transfer (CT) transition at around 600 nm in the absorption spectrum and a uniquely small A || in its electron paramagnetic resonance (EPR) spectrum. This is the site of substrate oxidation, and from table 1, the MCOs can be divided into two classes depending upon the identity of the substrate. For enzymes such as laccase 3 and ascorbate oxidase, 4 redox active organic molecules which can interact weakly with the enzyme provide the electrons. For MCOs like Fet3p 5 and Ceruloplasmin, 6 the substrate is a metal ion (ferrous in these cases) which binds tightly to a substrate binding site. As shown in figure 1, these substrate binding sites are located near the His ligands of the T1 Cu center. The electron from substrate is first transferred to the T1 and then over >13Å through a Cys-His pathway to a trinuclear Cu cluster (TNC) where O 2 is reduced to water (vide infra). 7 We first consider the electron transfer (ET) pathways to the TNC. ET PathwaysHere we focus on the Fe(II) binding site of the enzyme Fet3p, which is involved in the uptake of iron by yeast. 8 (Studies on this enzyme were performed in collaboration with Prof. Dan Kosman and coworkers.) A variable-temperature, variable-field magnetic circular dichroism (VTVH MCD) methodology we developed in other studies was applied to probe this ferrous site. [9][10][11] From figure 2A dark blue, there is a characteristic feature at 8900 cm −1 in the MCD spectrum corresponding to Fe(II) binding with a high affinity (K B > 10 5 M −1 , from MCD titration studies) to a 6 coordinate site in the protein. 12 In the light blue spectrum this feature is eliminated and a peak at 9700 cm −1 , corresponding to aqueous Fe(II) (green) is observed when Zn(II) is first bound to the substrate site, inhibiting ferroxidase activity.From mutagenesis studies we have found that three carboxylates are involved...

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Magnetic circular dichroism spectroscopy as a probe of the geometric and electronic structure of non-heme ferrous enzymes

Cited by 217 publications

References 164 publications

Spectroscopic and electronic structure studies of aromatic electrophilic attack and hydrogen-atom abstraction by non-heme iron enzymes

Spectroscopic and electronic structure studies of aromatic electrophilic attack and hydrogen-atom abstraction by non-heme iron enzymes

Magnetic Circular Dichroism of Paramagnetic Species

O2 Reduction to H2O by the multicopper oxidases

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