Diiron-oxo Proteins Research Articles

ChemInform Abstract: Artificial Diiron Enzymes with a de novo Designed Four‐Helix Bundle Structure

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Artificial Diiron Enzymes with a De Novo Designed Four‐Helix Bundle Structure

AbstractInvited for the cover of this issue is the group of Angela Lombardi at the University of Napoli “Federico II”, Italy. The cover image shows a construction site where workers are building up a four‐helix bundle scaffold. Fine‐tuning of the metal binding site, which is the heart of the construction, enables a specific function, such as oxidative chemistry. This approach has been successfully employed to develop diiron–oxo protein models.

Artificial Diiron Enzymes with a De Novo Designed Four-Helix Bundle Structure.

A single polypeptide chain may provide an astronomical number of conformers. Nature selected only a trivial number of them through evolution, composing an alphabet of scaffolds, that can afford the complete set of chemical reactions needed to support life. These structural templates are so stable that they allow several mutations without disruption of the global folding, even having the ability to bind several exogenous cofactors. With this perspective, metal cofactors play a crucial role in the regulation and catalysis of several processes. Nature is able to modulate the chemistry of metals, adopting only a few ligands and slightly different geometries. Several scaffolds and metal-binding motifs are representing the focus of intense interest in the literature. This review discusses the widespread four-helix bundle fold, adopted as a scaffold for metal binding sites in the context of de novo protein design to obtain basic biochemical components for biosensing or catalysis. In particular, we describe the rational refinement of structure/function in diiron-oxo protein models from the due ferri (DF) family. The DF proteins were developed by us through an iterative process of design and rigorous characterization, which has allowed a shift from structural to functional models. The examples reported herein demonstrate the importance of the synergic application of de novo design methods as well as spectroscopic and structural characterization to optimize the catalytic performance of artificial enzymes.

An artificial di-iron oxo-protein with phenol oxidase activity

Here we report the de novo design and NMR structure of a four-helical bundle di-iron protein with phenol oxidase activity. The introduction of the cofactor-binding and phenol-binding sites required the incorporation of residues that were detrimental to the free energy of folding of the protein. Sufficient stability was, however, obtained by optimizing the sequence of a loop distant from the active site.

Diiron-containing metalloproteins: Developing functional models

Abstract A major objective in protein science is the design of enzymes with novel catalytic activities that are tailored to specific applications. Such enzymes may have great potential in biocatalysis and biosensor technology, such as in degradation of pollutants and biomass, and in drug and food processing. To reach this objective, investigations into the basic biochemical functioning of metalloproteins are still required. In this perspective, metalloprotein design provides a powerful approach first to contribute to a more comprehensive understanding of the way metalloproteins function in biology, with the ultimate goal of developing novel biocatalysts and sensing devices. Metalloprotein mimetics have been developed through the introduction of novel metal-binding sites into naturally occurring proteins as well as through de novo protein design. We have approached the challenge of reproducing metalloprotein active sites by using a miniaturization process. We centered our attention on iron-containing proteins, and we developed models for heme proteins and diiron–oxo proteins. In this paper we summarize the results we obtained on the design, structural, and functional properties of DFs, a family of artificial diiron proteins.

Density functional theory of spin-coupled models for diiron-oxo proteins: Effects of oxo and hydroxo bridging on geometry, electronic structure, and magnetism

We have performed a comprehensive study of the electronic structure and magnetic properties of structurally characterized models for diiron-oxo proteins. Results from Kohn–Sham density functional theory show that two complexes, with formula Fe2(μ-O)(μ-O2CCH3)2(HBpz3)2 and [Fe2(μ-OH)(μ-O2CCH3)2(HBpz3)2]+, are strongly and weakly antiferromagnetically coupled, respectively, in agreement with experiment. The physical origin of the stronger and weaker exchange typically measured for oxo- and hydroxo-bridged diiron complexes, respectively, has been elucidated. The main superexchange pathways giving rise to molecular antiferromagnetism in both complexes have been identified. The dominant pathway in the oxo-bridged complex, Fe1(dxz):μ-O(px):Fe2(dxz), was formed by π interactions whereas that of the hydroxo-bridged, Fe1(dz2):μ-OH(p∥):Fe2(dz2), was formed by σ interactions. We also found a pathway mediated by the bridging acetates, Fe1(dx2−y2):bis(μ-acetato):Fe2(dx2−y2), which induces weak antiferromagnetism in the oxo-bridged complex but is significantly more important in the hydroxo-bridged complex. The antiferromagnetic exchange constants that parameterize the Heisenberg Hamiltonian H=JS1⋅S2 have been predicted for both, strongly and weakly, coupled complexes. Overall, the signs, trends, and magnitudes of the theoretical values (Jμ-Ocalc=+152.7 cm−1, Jμ-OHcalc=+23.3 cm−1) were in excellent agreement with experiment. The geometries of the complete molecular structures have been optimized in C2v symmetry and used to calculate molecular properties such as atomic charges and spin densities. The electronic configurations (Fe:4s0.293d5.93,μ-O:2s1.922p4.99;Fe:4s0.303d5.82,μ-OH:2s1.822p5.25,H:1s0.51) of the respective binuclear cores revealed relatively high occupancies for the nominally ferric ions, thus reflecting a donating character of their immediate N3O3 coordination. In addition, the diiron-oxo protein hemerythrin has been discussed. Theoretical and structural considerations indicated that the oxo-bridged diferric complex considered herein models extremely well the antiferromagnetic behavior of azidomet- and azidometmyo-hemerythrin. Finally, the magnetic behavior of closely related oxo-bridged diferric and hydroxo-bridged diferrous complexes containing Me3TACN capping ligands has been explained in light of the results presented in this work.

Synthesis, crystal structure of diferrous complex and its reactivity with dioxygen

The dinuclear complex of [Fe2L{O2P(OPh)2}](ClO4)2· Et2O(1) (where L represents the dinucleating ligand N-Et-HPTB, anion of N, N, N’, N’-tetrakis (N-ethyl-2-benzimidazolyemethyl)2-hydroxy-1, 3-diamino-propane) has been synthesized and crystallizes in the triclinic space group P1 with cell constants a = 1.526(3) nm, b = 1.259 8(3) nm, c = 1.563 0(3)° nm, α = 94.41(3)°, β = 115.31(3)°, γ = 99.90(3)°, V = 3.267(1) nm3, z = 2, R = 0.084 7 and Rw = 0.177 8. The Fe(II) sites are bridged by an alkoxide of the dinucleating ligand and a phosphate, affording a diiron core with an Fe-μ-O-Fe angle of 131.20(3)° and an Fe-Fe distance of 0.364 9 nm. Both Fe(II) centers have trigonal bipyramidal geometry. Dioxygen adduct (1/O2) forms upon exposure of the diferrous complex to O2 at low temperature (?60°C). The 1/O2 adduct is stable at ? 60°C but decomposes upon warming. The adduct exhibits visible absorption maximum near 606 nm and resonance Raman features at 478 cm-1 (γFe-O) and 897 cm-1 (γO-O), and the latter is the characteristic of a μ-1,2-peroxo species, indicating that dioxygen adducts can serve as models for the putative oxygenated intermediate of some non-heme diiron-oxo proteins.

O2 Activation and Aromatic Hydroxylation Performed by Diiron Complexes

Chemical models of active sites of diiron oxo proteins have been synthesized. The polydendate ligands are EDTA derivatives which provide a balanced supply of nitrogen atoms and carboxylate groups t...

Structural similarity and functional diversity in diiron-oxo proteins

Diiron-oxo proteins currently represent one of the most rapidly developing areas of bioinorganic chemistry. All of these proteins contain a four-helix bundle protein fold surrounding a (μ-carboxylato)diiron core, and most, if not all, of the diiron(II) sites appear to react with O2 as part of their functional processes. Despite these common characteristics, an emerging functional diversity is one of the most striking aspects of this class of proteins. X-ray crystal structures of diiron(II) sites are now available for four of these proteins: hemerythrin (Hr), the hydroxylase protein of methane monooxygenase (MMOH), the R2 protein of Escherichia coli ribonucleotide reductase (RNR-R2), and a plant acyl-carrier protein Δ9-desaturase. The structure of the diiron(II) site in Hr, the sole O2 carrier in the group, is clearly distinct from the other three, whose function is oxygen activation. The Hr diiron site is more histidine rich, and the oxygen-activating diiron sites contain a pair of (D/E)X30–37EX2H ligand sequence motifs, which is clearly not found in Hr. The Hr diiron site apparently permits only terminal O2 coordination to a single iron, whereas the oxygen-activating diiron(II) centers present open or labile coordination sites on both irons of the center, and show a much greater coordinative flexibility upon oxidation to the diiron(III) state. Intermediates at the formal FeIIIFeIII and FeIVFeIV oxidation levels for MMOH and formal FeIIIFeIV oxidation level for RNR-R2 have been identified during reactions of the diiron(II) sites with O2. An [Fe2(μ-O)2]4+, 3+ "diamond core" structure has been proposed for the latter two oxidation levels. The intermediate at the FeIIIFeIV oxidation level in RNR-R2 is kinetically competent to generate a stable, functionally essential tyrosyl radical. The FeIVFeIV oxidation level is presumed to effect hydroxylation of hydrocarbons in MMOH, but the mechanism of this hydroxylation, particularly the involvement of discrete radicals, is currently controversial. The biological function of diiron sites in three members of this class, rubrerythrin, ferritin and bacterioferritin, remains enigmatic.

Dioxgen binding to diferrous centers. Models for diiron-oxo proteins

Dioxgen binding to diferrous centers. Models for diiron-oxo proteins

Insight into the g .apprxeq. 16 EPR signals of reduced diiron-oxo proteins. Structure and properties of [FeII2BPMP{O2P(OC6H5)2}2]Cl

The diiron complexes [Fe II 2 BPMP{O 2 P(OC 6 H 5 ) 2 } 2 ]X (1 (X=Cl), 2 (X=BF 4 ), 3 (X=X=BPh 4 )) and [Fe II 2 BPCP-(O 2 CC 2 H 5 ) 2 ]BPh 4 (4), where BPMP is the anion of 2,6-bis[(bis(2-pyridylmethyl)amino)methyl]-4-methylphenol and BPCP is the anion of 2,6-bis[(bis(2-pyridylmethyl)amino)methyl]-4-chlorophenol, have been synthesized to provide insight into the integer-spin EPR signals found in the diferrous forms of diiron-oxo proteins

Dioxygen binding to diferrous centers. Models for diiron-oxo proteins

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTDioxygen binding to diferrous centers. Models for diiron-oxo proteinsYanhong Dong, Stephane Menage, Bridget A. Brennan, Timothy E. Elgren, Ho G. Jang, Linda L. Pearce, and Lawrence Que Jr.Cite this: J. Am. Chem. Soc. 1993, 115, 5, 1851–1859Publication Date (Print):March 1, 1993Publication History Published online1 May 2002Published inissue 1 March 1993https://doi.org/10.1021/ja00058a033RIGHTS & PERMISSIONSArticle Views1076Altmetric-Citations121LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (1 MB) Get e-AlertsSupporting Info (2)»Supporting Information Supporting Information Get e-Alerts

2D NMR studies of paramagnetic diiron complexes

Diiron complexes of dinucleating ligands have been demonstrated to be good structural and spectroscopic models for the diiron-oxo proteins. Although such complexes show relatively sharp isotropically shifted 1 H NMR resonances due to their fast electronic relaxation rates, detailed assignment of their isotropically shifted 1 H NMR resonances has thus far not been achieved due to the difficulty in esablishing bond connectivities on paramagnetic complexes

Effects of a bridging dicarboxylate ligand on the synthesis and physical properties of (.mu.-oxo)bis(.mu.-carboxylato)diiron(III) complexes

The molecule m-phenylenedipropionic acid (H 2 MPDP), in which two carboxylates are part of the same bridging unit, has been used in the synthesis of diiron(III) oxo protein analogues containing the (μ-oxo)bis(μ-carboxylato)diiron(III) core. The dicarboxylate ligand MPDP 2− facilitates the self-assembly synthesis of [Fe 2 O(MPDP){HB(pz) 3 } 2 ] and [Fe 2 O(MPDP)B 2 Cl 2 ], complexes having terminal bidentate, B, dipyridyl or bis(1-methylimidazol-2-yl)phenylmethoxymethane (BIPhMe, a polyimidazole nitrogen donor), and monodentate chloride ligands

Models for diiron-oxo proteins: the peroxide adduct of Fe2(HPTB)(OH)(NO3)4

The complex of Fe(NO 3 ) 3 and N,N,N',N'-tetrakis(2-benzimidazolylmethyl)-2-hydroxy-1,3-diaminopropane (HPTB) is reformulated as [Fe 2 (HPTB)(μ-OH)(NO 3 ) 2 ](NO 3 ) 2 on the basis of 1 H NMR, EXAFS, X-ray diffraction, and conductivity data. This complex reacts with hydrogen peroxide to form a 1:1 adduct with a new charge-transfer band at 600 nm