Biosynthesis of a new skyllamycin in : a cytochrome P450 forms an epoxide in the cinnamoyl chain.

Activation of a silent gene cluster in leads to synthesis of a cinnamoyl-containing non-ribosomal peptide (CCNP) that is related to skyllamycins. This novel CCNP was isolated and its structure was interrogated using mass spectrometry and nuclear magnetic resonance spectroscopy. The isolated compound is an oxidised skyllamycin A in which an additional oxygen atom is incorporated in the cinnamoyl side-chain in the form of an epoxide.

Enhancing ferryl accumulation in HO-dependent cytochrome P450s.

A facile strategy is presented to enhance the accumulation of ferryl (iron(IV)-oxo) species in HO dependent cytochrome P450s (CYPs) of the CYP152 family. We report the characterization of a highly chemoselective CYP decarboxylase from Staphylococcus aureus (OleT) that is soluble at high concentrations. Examination of OleT Compound I (CpdI) accumulation with a variety of fatty acid substrates reveals a dependence on resting spin-state equilibrium.

Experimental and Theoretical Examination of the Kinetic Isotope Effect in Cytochrome P450 Decarboxylase OleT.

Using a combination of experimental studies, theory, simulation, and modeling, we investigate the hydrogen atom transfer (HAT) reaction by the high-valent ferryl cytochrome P450 (CYP) intermediate known as Compound I, a species that is central to innumerable and important detoxification and biosynthetic reactions. The P450 decarboxylase known as OleT converts fatty acids, a sustainable biological feedstock, into terminal alkenes and thus is of high interest as a potential means to produce fungible biofuels. Previous experimental work has established the intermediacy of Compound I in the C─C scission reaction catalyzed by OleT and an unprecedented ability to monitor the HAT process in the presence of bound fatty acid substrates.

Structural analysis of P450 AmphL from Streptomyces nodosus provides insights into substrate selectivity of polyene macrolide antibiotic biosynthetic P450s.

AmphL is a cytochrome P450 enzyme that catalyzes the C8 oxidation of 8-deoxyamphotericin B to the polyene macrolide antibiotic, amphotericin B. To understand this substrate selectivity, we solved the crystal structure of AmphL to a resolution of 2.0 Å in complex with amphotericin B and performed molecular dynamics (MD) simulations.

Proton Relay Network in the Bacterial P450s: CYP101A1 and CYP101D1.

Cytochrome P450s are among nature's most powerful catalysts. Their ability to activate molecular dioxygen to form high-valent ferryl intermediates (Compounds I and II) enables a wide array of chemistries ranging from simple epoxidations to more complicated C-H bond oxidations. Oxygen activation is achieved by reduction of the ferrous dioxygen complex, which requires the transfer of an electron from a redox partner and subsequent double protonation to yield a water molecule and a ferryl porphyrin π-cation radical (Compound I).

Unexpected Differences between Two Closely Related Bacterial P450 Camphor Monooxygenases.

The bacterial cytochrome P450cam catalyzes the oxidation of camphor to 5--hydroxycamphor as the first step in the oxidative assimilation of camphor as a carbon/energy source. CYP101D1 is another bacterial P450 that catalyzes the same reaction. A third P450 (P450tcu) has recently been discovered that has ≈86% sequence identity to P450cam as well as very similar enzymatic properties.

A Distal Loop Controls Product Release and Chemo- and Regioselectivity in Cytochrome P450 Decarboxylases.

Cytochrome P450 OleT utilizes hydrogen peroxide (HO) to catalyze the decarboxylation or hydroxylation of fatty acid (FA) substrates. Both reactions are initiated through the abstraction of a substrate hydrogen atom by the high-valent iron-oxo intermediate known as Compound I. Here, we specifically probe the influence of substrate coordination on OleT reaction partitioning through the combined use of fluorescent and electron paramagnetic resonance (EPR)-active FA probes and mutagenesis of a structurally disordered F-G loop that is distal from the heme-iron active site.

On the Structure and Reaction Mechanism of Human Acireductone Dioxygenase.

Acireductone dioxygenase (ARD) is an intriguing enzyme from the methionine salvage pathway that is capable of catalysing two different oxidation reactions with the same substrate depending on the type of the metal ion in the active site. To date, the structural information regarding the ARD-acireductone complex is limited and possible reaction mechanisms are still under debate. The results of joint experimental and computational studies undertaken to advance knowledge about ARD are reported.

Activation of C-H Bonds of Alkyl- and Arylnitriles by the TaCl-PPh Lewis Pair.

A new pathway of activation of C-H bonds of alkyl- and arylnitriles by a cooperative action of TaCl and PPh under mild conditions is reported. Coordination of nitriles to the highly Lewis acidic Ta(V) center resulted in an activation of their aliphatic and aromatic C-H bonds, allowing nucleophilic attack and deprotonation by the relatively weak base PPh. The propensity of Ta(V) to form multiple bonds to nitrogen-containing ligands is an important driving force of the reaction as it led to a sequence of bond rearrangements and the emergence of, in the case of benzonitrile, a zwitterionic enediimido complex of Ta(V) through C═C double bond formation between two activated nitrile fragments.

The Enigmatic P450 Decarboxylase OleT Is Capable of, but Evolved To Frustrate, Oxygen Rebound Chemistry.

OleT is a cytochrome P450 enzyme that catalyzes the removal of carbon dioxide from variable chain length fatty acids to form 1-alkenes. In this work, we examine the binding and metabolic profile of OleT with shorter chain length (n ≤ 12) fatty acids that can form liquid transportation fuels. Transient kinetics and product analyses confirm that OleT capably activates hydrogen peroxide with shorter substrates to form the high-valent intermediate Compound I and largely performs C-C bond scission.

Divergent mechanisms of iron-containing enzymes for hydrocarbon biosynthesis.

Increasing levels of energy consumption, dwindling resources, and environmental considerations have served as compelling motivations to explore renewable alternatives to petroleum-based fuels, including enzymatic routes for hydrocarbon synthesis. Phylogenetically diverse species have long been recognized to produce hydrocarbons, but many of the enzymes responsible have been identified within the past decade. The enzymatic conversion of C chain length fatty aldehydes (or acids) to C hydrocarbons, alkanes or alkenes, involves a C-C scission reaction.

Mixed regiospecificity compromises alkene synthesis by a cytochrome P450 peroxygenase from Methylobacterium populi.

Intensive interest has focused on enzymes that are capable of synthesizing hydrocarbons, alkenes and alkanes, for sustainable fuel production. A recently described cytochrome P450 (OleTJE) from the CYP152 family catalyzes an unusual carbon-carbon scission reaction, transforming Cn fatty acids to Cn-1 1-alkenes. Here, we show that a second CYP152, CYP-MP from Methylobacterium populi ATCC BAA 705, also catalyzes oxidative substrate decarboxylation.