Bioinspired molecular catalysts for homogenous electrochemical activation of dioxygen

International audienceO2, which is abundant and environmentally benign, is the ideal green oxidant for oxidation reactions, which are key transformations in the chemical industry. Still, O2 needs to be activated, and this can be achieved through the so-called reductive activation of the O2 paradigm. Taking inspiration from metalloenzymes, where a non-noble redox active metal (iron, copper) controls the partial reduction of O2 via electron and proton transfers, metal-based synthetic systems can be developed to reproduce oxygenase activity. In the present article, we focus on fundamental aspects that serve as support for the development of 2-electron activation of O2 and generation of highly oxidizing metal-oxo species, thus paving the road for the development of electrocatalytic systems for organic substrate oxygenation. Scarce examples known in the literature capable of such reactivity and possible future developments are described

Hijacking Chemical Reactions of P450 Enzymes for Altered Chemical Reactions and Asymmetric Synthesis

Cytochrome P450s are heme-containing enzymes capable of the oxidative transformation of a wide range of organic substrates. A protein scaffold that coordinates the heme iron, and the catalytic pocket residues, together, determine the reaction selectivity and regio- and stereo-selectivity of the P450 enzymes. Different substrates also affect the properties of P450s by binding to its catalytic pocket. Modulating the redox potential of the heme by substituting iron-coordinating residues changes the chemical reaction, the type of cofactor requirement, and the stereoselectivity of P450s. Around hundreds of P450s are experimentally characterized, therefore, a mechanistic understanding of the factors affecting their catalysis is increasingly vital in the age of synthetic biology and biotechnology. Engineering P450s can enable them to catalyze a variety of chemical reactions viz. oxygenation, peroxygenation, cyclopropanation, epoxidation, nitration, etc., to synthesize high-value chiral organic molecules with exceptionally high stereo- and regioselectivity and catalytic efficiency. This review will focus on recent studies of the mechanistic understandings of the modulation of heme redox potential in the engineered P450 variants, and the effect of small decoy molecules, dual function small molecules, and substrate mimetics on the type of chemical reaction and the catalytic cycle of the P450 enzymes

Bioinorganic Chemistry

This book covers material that could be included in a one-quarter or one-semester course in bioinorganic chemistry for graduate students and advanced undergraduate students in chemistry or biochemistry. We believe that such a course should provide students with the background required to follow the research literature in the field. The topics were chosen to represent those areas of bioinorganic chemistry that are mature enough for textbook presentation. Although each chapter presents material at a more advanced level than that of bioinorganic textbooks published previously, the chapters are not specialized review articles. What we have attempted to do in each chapter is to teach the underlying principles of bioinorganic chemistry as well as outlining the state of knowledge in selected areas. We have chosen not to include abbreviated summaries of the inorganic chemistry, biochemistry, and spectroscopy that students may need as background in order to master the material presented. We instead assume that the instructor using this book will assign reading from relevant sources that is appropriate to the background of the students taking the course. For the convenience of the instructors, students, and other readers of this book, we have included an appendix that lists references to reviews of the research literature that we have found to be particularly useful in our courses on bioinorganic chemistry

Oxygen Activation and Radical Transformations in Heme Proteins and Metalloporphyrins

As a result of the adaptation of life to an aerobic environment, nature has evolved a panoply of metalloproteins for oxidative metabolism and protection against reactive oxygen species. Despite the diverse structures and functions of these proteins, they share common mechanistic grounds. An open-shell transition metal like iron or copper is employed to interact with O_2 and its derived intermediates such as hydrogen peroxide to afford a variety of metal–oxygen intermediates. These reactive intermediates, including metal-superoxo, -(hydro)peroxo, and high-valent metal–oxo species, are the basis for the various biological functions of O_2-utilizing metalloproteins. Collectively, these processes are called oxygen activation. Much of our understanding of the reactivity of these reactive intermediates has come from the study of heme-containing proteins and related metalloporphyrin compounds. These studies not only have deepened our understanding of various functions of heme proteins, such as O2 storage and transport, degradation of reactive oxygen species, redox signaling, and biological oxygenation, etc., but also have driven the development of bioinorganic chemistry and biomimetic catalysis. In this review, we survey the range of O_2 activation processes mediated by heme proteins and model compounds with a focus on recent progress in the characterization and reactivity of important iron–oxygen intermediates. Representative reactions initiated by these reactive intermediates as well as some context from prior decades will also be presented. We will discuss the fundamental mechanistic features of these transformations and delineate the underlying structural and electronic factors that contribute to the spectrum of reactivities that has been observed in nature as well as those that have been invented using these paradigms. Given the recent developments in biocatalysis for non-natural chemistries and the renaissance of radical chemistry in organic synthesis, we envision that new enzymatic and synthetic transformations will emerge based on the radical processes mediated by metalloproteins and their synthetic analogs

Functional Characterization of Synthetic Metalloporphyrin-Containing Enzymes

Over the last decades, the analysis of the catalytic properties of metalloenzymes has received increasing attention, because of the variety of complex chemical transformations that these molecules are able to perform. The excellent catalytic potential of metalloenzymes derives from their aptitude to easily form highly reactive intermediates, resulting from the stabilization of metal ion oxidation states. In the effort to resemble (or even enhance) the catalytic properties of natural metalloenzymes while improving their chemical stability, a wide variety of bioinspired systems have been devised, obtained modifying either the metal cofactor or the protein matrix. In this context, the development of a novel artificial heme-enzyme belonging to the Mimochrome family (Mimochrome VIa or MC6a) and the evaluation of its catalytic potential have been explored during this PhD project. In early studies, the design, synthesis and characterization of the FeIII-containing MC6a (FeIII-MC6a) have been performed. Compared to the best candidate among previous mimochrome members, FeIII-MC6a acted as an even more efficient peroxidase-like catalyst, since it was able to oxidize ABTS (taken as model substrate) with a much higher turnover number (from 5900 to 14000) and turnover frequency (from 2300 to 5900 s-1). This feature has been correlated with the presence of two Aib residues in place of Gln3 e Ser7 on the (D) chain. Indeed, this substitution has been found to significantly increase the conformational rigidity of the peptide chain, contributing to a higher preorganization of the molecule toward protein folding. Furthermore, the catalyst was endowed with a higher robustness than that of all previous mimochrome members (up to a two-fold increase), although at the cost of a minor affinity toward H2O2 (up to a three-fold increase of KM value). In this case, it has been conjectured that the larger hydrophobic character of the (D) chain due to the presence of Aib residues may have resulted into additional interactions between the peptide chain and the porphyrin ring, thereby providing a more compact structure, which protects the metal cofactor from H2O2-mediated bleaching. In subsequent studies, the analysis of the reactivity of a FeIII-MC6a derivative containing a CoIII ion (Co-MC6a) has been conducted. These studies, which were mainly carried out under the supervision of Prof. K. L. Bren (University of Rochester, NY), were aimed to investigate the potential use of MC6a in hydrogen evolution reactions (HERs), in view of a potential employ of our catalyst in water splitting approaches. In this case, Co-MC6a is intended to represent an alternative to natural hydrogenases, whose use as catalysts for HERs is limited by their sensitivity to molecular oxygen. Co-MC6a demonstrated to be a convenient electrocatalyst in HERs. Differently from most natural enzymes or synthetic organometallic biomimetics, it acted as a water-soluble catalyst, working in neutral water and in the presence of molecular oxygen. In addition, compared to previous Co-porphyrin based synthetic enzymes (e.g. Co-MP11), Co-MC6a worked with similar turnover frequency and overpotential values but with much higher turnover numbers (up to 300000), retaining its activity even after several hours. Interestingly, the comparative analysis of Co-MP11 with Co-MC6a enabled to correlate the overpotential value in HERs with the enzyme folding. In addition, preliminary SAR studies have been undertaken, in order to identify the existing correlation between coordination shell of our enzyme and its efficacy in HERs. The result of this study will allow exploring the chemical space enabling further improvement in the enzyme-like properties of our catalyst. Overall, the results obtained during this PhD thesis represent a significant improvement in our knowledge of peptide-based artificial metalloenzymes, as they have contributed to provide the most advanced candidate with catalytic potential. Given the ease in having access to these molecules and their mutants, future endeavours will be addressed to an in-depth study of the structural requirements for enzymatic activity, in order to develop even higher performing catalysts than natural metalloenzymes. Furthermore, given the possibility to easily change the metal cofactors as well as their coordination spheres, efforts will be focused in widening the repertoire of reactions and thereby the potential applications of these enzymes

Nature’s Machinery, Repurposed: Expanding the Repertoire of Iron-Dependent Oxygenases

Iron is an especially important redox-active cofactor in biology because of its ability to mediate reactions with atmospheric O₂. Iron-dependent oxygenases exploit this earth-abundant transition metal for the insertion of oxygen atoms into organic compounds. Throughout the astounding diversity of transformations catalyzed by these enzymes, the protein framework directs reactive intermediates toward the precise formation of products, which, in many cases, necessitates the cleavage of strong C–H bonds. In recent years, members of several iron-dependent oxygenase families have been engineered for new-to-nature transformations that offer advantages over conventional synthetic methods. In this Perspective, we first explore what is known about the reactivity of heme-dependent cytochrome P450 oxygenases and nonheme iron-dependent oxygenases bearing the 2-His-1-carboxylate facial triad by reviewing mechanistic studies with an emphasis on how the protein scaffold maximizes the catalytic potential of the iron-heme and iron cofactors. We then review how these cofactors have been repurposed for abiological transformations by engineering the protein frameworks of these enzymes. Finally, we discuss contemporary challenges associated with engineering these platforms and comment on their roles in biocatalysis moving forward

Nature’s Machinery, Repurposed: Expanding the Repertoire of Iron-Dependent Oxygenases

Iron is an especially important redox-active cofactor in biology because of its ability to mediate reactions with atmospheric O₂. Iron-dependent oxygenases exploit this earth-abundant transition metal for the insertion of oxygen atoms into organic compounds. Throughout the astounding diversity of transformations catalyzed by these enzymes, the protein framework directs reactive intermediates toward the precise formation of products, which, in many cases, necessitates the cleavage of strong C–H bonds. In recent years, members of several iron-dependent oxygenase families have been engineered for new-to-nature transformations that offer advantages over conventional synthetic methods. In this Perspective, we first explore what is known about the reactivity of heme-dependent cytochrome P450 oxygenases and nonheme iron-dependent oxygenases bearing the 2-His-1-carboxylate facial triad by reviewing mechanistic studies with an emphasis on how the protein scaffold maximizes the catalytic potential of the iron-heme and iron cofactors. We then review how these cofactors have been repurposed for abiological transformations by engineering the protein frameworks of these enzymes. Finally, we discuss contemporary challenges associated with engineering these platforms and comment on their roles in biocatalysis moving forward

GENERATION, CHARACTERIZATION AND REACTIVITY OF COBALT DIAMOND CORE AND COBALT PEROXO COMPLEXES

The development of efficient and low-cost technologies that convert hydrocarbons, including methane, to liquid fuels through controlled functionalization of its inert C–H bond is a fundamental challenge. Also, the activation of carbon−hydrogen (C−H) bonds is the first step of functionalizing inert hydrocarbons. This transformation is a key step in many biological and synthetic processes. One representative example inspired by nature is the metalloenzyme called soluble methane monooxygenase (sMMO), a nonheme dinuclear iron-dependent enzyme that catalyzes the hydroxylation of the strong C–H bond of methane (bond dissociation energy BDE = 105 kcal/mol) using O2 as the oxidant. The catalytic cycle of sMMO has been extensively studied over decades, and features a highvalent bis-μ-oxo FeIV2(μ-O)2 “diamond core” intermediate called Q as the active oxidant for C–H bond activation. This research focuses on the study of an unprecedented highvalent CoIII,IV2(μ-O)2 complex supported by neutral tetradentate tris(2- pyridylmethyl)amine (TPA) ligand by one-electron oxidation of its CoIII2(μ-O)2 precursor. This new complex can activate C−H bonds 3−5 orders of magnitude faster than its iron and manganese counterparts, and represents the most reactive synthetic model for the sMMO enzymatic intermediate. This study expands the understanding of base metal complexes for C−H bond activation and serves as motivation to design C−H activation methods inspired by nature. Chapter 1 provides the introduction of C-H bond hydroxylation mechanism and the biological background that initially inspired this project. In Chapter 2, we reported the characterization of cobalt diamond core complexes supported by TPA and related ligands. In Chapter 3, we studied the reactivity of those complexes. In Chapter 4, we discovered that the open core species provides an excellent strategy to achieve substrate specificity and to be applied in the deaminative C(sp3)-N bond activation. Chapter 5 describes a proposed monomer [Co(III)(TPA)(O2)]+ species and its nucleophilic reactivity. Chapter 6 lays out the overall conclusions and points out a few future directions as the prospective scope of the entire project

HEME-IRON AND HEME/COPPER OXYGEN CHEMISTRY OF METALLOENZYME ACTIVE SITES WITH BIO-INSPIRED SYNTHETIC MODEL SYSTEM

The activation and reduction of dioxygen are the most important process in biological respiration and in energy conversion systems such as fuel cells which harness clean and effective electrical power generated from chemical fuels that use O2 as an electron/cation acceptor. The stepwise reduction of dioxygen to water is highly exothermic transformation which is substantial for aerobic respiration. The metal-bound reduced-O2 intermediates are formed in the course of reductive O‒O bond activation. Establishing redox and thermodynamic relationships between metal-oxy species and its reduced (and protonated) derivatives is critically important for a full understanding of (bio)chemical processes involving metalloenzyme mediated dioxygen processing. Biomimetic synthetic model chemistry is a powerful tool to better understand fundamental elements of metalloprotein electronics, functions, and selectivity/reactivity. In chapter 1, an overview of reductive dioxygen activation by heme and heme-peroxide-copper based systems is provided. It also includes important intermediates in the catalytic reduction of dioxygen by metalloenzymes. Chapter 2 presents a new iron-porphyrinate complex, which employs a tridentate ligand that incorporated a histamine moiety appended to the periphery of a fluorinated tetraphenylporphyrin and the reactivity of various reduced heme compounds, toward 2,6-dimethyl-phenyl isocyanide (DIMPI) and nitric oxide (NO). The generated DIMPI-FeII and NO-FeII complexes were characterized by UV-vis, IR, NMR, and EPR spectroscopies. In chapter 3, the dioxygen reactivity of reduced heme, which is an advanced cytochrome c oxidase (CcO) active site model system as a binucleating ligand, is reported in the absence and the presence of copper ion. This work represents that the process of oxygenation of the iron complex in the absence and presence of copper ion matches well with proposed CcO catalytic cycle, i.e., Cu-independent generation of heme superoxide and subsequent formation of peroxide moiety with copper ion. Chapter 4 discusses the stepwise reduction and protonation of a ferric heme superoxide complex and the first example of experimentally determined thermodynamics (reduction potential and pKa). With these measure thermodynamic parameters, the OO–H bond dissociation free energy (BDFE) was calculated employing thermodynamic square scheme and Bordwell equation. The determined BDFE value was confirmed by the oxidizing capability of ferric heme superoxide species via hydrogen atom transfer (HAT). In chapter 5, thermodynamic comparisons for O2-derived iron-porphyrinate interrelated ferric superoxide, peroxide and hydroperoxide complexes in the presence and absence of an appended imidazolyl axial base are described. Also, the reactivity results of superoxide species with/without axial ligand with external substrates reveal their oxidizing capability, and these observations corroborate the new thermodynamic results presented here