Two decades have passed since the discovery in liver microsomes of a haemprotein that forms a reduced-CO complex with the absorptive maximum of the Soret at 450 nm (Klingenberg, 1958; Garfinkel, 1958) and the identification of this protein as a new cytochrome: pigment cytochrome, P-450 (Omura and Sato, 1962, 1964a). In the intervening years, the study of cytochrome P-450 dependent monoxygenases has expanded exponentially. From the first crude attempts to solubilise a P-450 (Omura and Sato, 1963, 1964b) to the determination of the primary, secondary, and tertiary structure of cytochrome P-450cam by amino acid sequencing (Haniu et al., 1982a,b) and x-ray crystallography (Poulos et al., 1984) our understanding of this unique family of proteins has been advancing on all fronts. Since, perhaps, the greatest understanding of the structure and mechanism of P-450s has come from concentrated study of P-450cam of the Pseudomonas putida camphor-5-exo-hydroxylase, this review will concentrate on findings with P-450cam; attention will be drawn to parallel and contrasting examples from other P-450s as appropriate.

Structure and Chemistry of Cytochrome P450

Globally increasing energy demands and environmental concerns related to the use of fossil fuels have stimulated extensive research to identify new energy systems and economies that are sustainable, clean, low cost, and environmentally benign. Hydrogen generation from solar-driven water splitting is a promising strategy to store solar energy in chemical bonds. The subsequent combustion of hydrogen in fuel cells produces electric energy, and the only exhaust is water. These two reactions compose an ideal process to provide clean and sustainable energy. In such a process, a hydrogen evolution reaction (HER), an oxygen evolution reaction (OER) during water splitting, and an oxygen reduction reaction (ORR) as a fuel cell cathodic reaction are key steps that affect the efficiency of the overall energy conversion. Catalysts play key roles in this process by improving the kinetics of these reactions. Porphyrin-based and corrole-based systems are versatile and can efficiently catalyze the ORR, OER, and HER. Becau...

Energy-Related Small Molecule Activation Reactions: Oxygen Reduction and Hydrogen and Oxygen Evolution Reactions Catalyzed by Porphyrin- and Corrole-Based Systems

The design of artificial catalysts able to compete with the catalytic proficiency of enzymes is an intense subject of research. Non-covalent interactions are thought to be involved in several properties of enzymatic catalysis, notably (i) the confinement of the substrates and the active site within a catalytic pocket, (ii) the creation of a hydrophobic pocket in water, (iii) self-replication properties and (iv) allosteric properties. The origins of the enhanced rates and high catalytic selectivities associated with these properties are still a matter of debate. Stabilisation of the transition state and favourable conformations of the active site and the product(s) are probably part of the answer. We present here artificial catalysts and biomacromolecule hybrid catalysts which constitute good models towards the development of truly competitive artificial enzymes.

/pdf/supramolecular-catalysis-part-2-artificial-enzyme-mimics-2h1esy0td8.pdf

Supramolecular catalysis. Part 2: artificial enzyme mimics

The incorporation of a synthetic, catalytically competent metallocofactor into a protein scaffold to generate an artificial metalloenzyme (ArM) has been explored since the late 1970’s. Progress in the ensuing years was limited by the tools available for both organometallic synthesis and protein engineering. Advances in both of these areas, combined with increased appreciation of the potential benefits of combining attractive features of both homogeneous catalysis and enzymatic catalysis, led to a resurgence of interest in ArMs starting in the early 2000’s. Perhaps the most intriguing of potential ArM properties is their ability to endow homogeneous catalysts with a genetic memory. Indeed, incorporating a homogeneous catalyst into a genetically encoded scaffold offers the opportunity to improve ArM performance by directed evolution. This capability could, in turn, lead to improvements in ArM efficiency similar to those obtained for natural enzymes, providing systems suitable for practical applications and ...

/pdf/artificial-metalloenzymes-reaction-scope-and-optimization-2fb45ld2i6.pdf

Artificial Metalloenzymes: Reaction Scope and Optimization Strategies.

1. Overview A 2. Protein Redesign B 2.1. Making Use of Native Proteins: Protein Redesign B 2.2. Protein Redesign Based on Functions C 2.2.1. Redesign of Zinc Finger Structural Sites C 2.2.2. Redesign of Zinc Hydrolytic Centers E 2.2.3. Redesign of Heme Centers J 2.2.4. Redesign of Nonheme Redox Centers M 2.2.5. Artificial Metalloenzymes for Regioand Enantioselective Catalysis AA 2.2.6. Redesigned Protein Assemblies as Nanoreactors AK 2.3. Summary AN 3. De Novo Design AN 3.1. A Minimalist Approach: Designing Proteins from Scratch AN 3.2. Interactions between De Novo Designed Peptides and Metal Ions AN 3.2.1. Heavy Metal Toxicity AO 3.2.2. De Novo Designed Metal Centers Based on β-Structures AT 3.2.3. Metal-Induced Protein Folding AV 3.3. De Novo Designed Functional Metalloproteins: The Grail Quest of Protein Design AX 3.3.1. De Novo Designed Hydrolytic Centers AX 3.3.2. De Novo Designed Electron Transfer Centers BB 3.3.3. Other Catalytic Centers BG 3.4. Summary BN 4. Perspective BO Author Information BO Corresponding Author BO Notes BO Biographies BO Abbreviations BR References BR

Protein design: toward functional metalloenzymes.

Peroxidase activity of a myoglobin reconstituted with a chemically modified heme 1 is reported. The heme 1 bearing a total of eight carboxylates bound to the terminal of propionate side chains is incorporated into apomyoglobin from horse heart to obtain a new reconstituted myoglobin, rMb(1), with a unique binding domain structure. The UV−vis, CD, and NMR spectra of rMb(1) are comparable with those of native myoglobin, nMb. The mixing of rMb(1) with hydrogen peroxide yields a peroxidase compound II-like species, rMb(1)-II, since the spectrum of rMb(1)-II is identical with that observed for nMb. Stoichiometric oxidation of several small molecules by rMb(1)-II, demonstrates the significant reactivity. (i) The oxidation of cationic substrate such as [Ru(NH3)6]2+ by rMb(1)-II is faster than that observed for oxoferryl species of nMb, nMb-II. (ii) Anionic substrates such as ferrocyanide are unsuitable for the oxidation by rMb(1)-II. (iii) Oxidations of catechol, hydroquinone, and guaiacol are dramatically enhan...

Peroxidase Activity of Myoglobin Is Enhanced by Chemical Mutation of Heme-Propionates

To modulate the physiological function of a hemoprotein, most approaches have been demonstrated by site-directed mutagenesis. Replacement of the native heme with an artificial prosthetic group is another way to modify a hemoprotein. However, an alternate method, mutation or heme reconstitution, does not always demonstrate sufficient improvement compared with the native heme enzyme. In the present study, to convert a simple oxygen storage hemoprotein, myoglobin, into an active peroxidase, we applied both methods at the same time. The native heme of myoglobin was replaced with a chemically modified heme 2 having two aromatic rings at the heme-propionate termini. The constructed myoglobins were examined for 2-methoxyphenol (guaiacol) oxidation in the presence of H2O2. Compared with native myoglobin, rMb(H64D·2) showed a 430-fold higher kcat/Km value, which is significantly higher than that of cytochrome c peroxidase and only 3-fold less than that of horseradish peroxidase. In addition, myoglobin-catalyzed de...

Hybridization of modified-heme reconstitution and distal histidine mutation to functionalize sperm whale myoglobin.

Photocatalytic hydrogen generation using a protein-coated photosensitizer with anionic patches and a monocationic electron mediator

The synthesis of the first fluorine-containing iron porphycenes, 2,7,12,17-tetraethyl-3,6,13,16-tetrakis(trifluoromethyl)porphycenatoiron(III) chloride [FePc(EtioCF3)]Cl and its μ-oxo dimer [FePc(E...

Tsutomu Ando

Papers

Peroxidase Activity of Myoglobin Is Enhanced by Chemical Mutation of Heme-Propionates

Hybridization of modified-heme reconstitution and distal histidine mutation to functionalize sperm whale myoglobin.

Photocatalytic hydrogen generation using a protein-coated photosensitizer with anionic patches and a monocationic electron mediator

Synthesis, characterization, and autoreduction of a highly electron-deficient porphycenatoiron(III) with trifluoromethyl substituents.

Introduction of a specific binding domain on myoglobin surface by new chemical modification