A glycyl radical enzyme enables hydrogen sulfide production by the human intestinal bacterium .

Hydrogen sulfide (HS) production in the intestinal microbiota has many contributions to human health and disease. An important source of HS in the human gut is anaerobic respiration of sulfite released from the abundant dietary and host-derived organic sulfonate substrate in the gut, taurine (2-aminoethanesulfonate). However, the enzymes that allow intestinal bacteria to access sulfite from taurine have not yet been identified.

Use of the dehydrophos biosynthetic enzymes to prepare antimicrobial analogs of alaphosphin.

The C-terminal domain of the dehydrophos biosynthetic enzyme DhpH (DhpH-C) catalyzes the condensation of Leu-tRNALeu with (R)-1-aminoethylphosphonate, the aminophosphonate analog of alanine called Ala(P). The product of this reaction, Leu-Ala(P), is a phosphonodipeptide, a class of compounds that have previously been investigated for use as clinical antibiotics. In this study, we show that DhpH-C is highly substrate tolerant and can condense various aminophosphonates (Gly(P), Ser(P), Val(P), 1-amino-propylphosphonate, and phenylglycine(P)) to Leu.

Structural basis for methylphosphonate biosynthesis.

Methylphosphonate synthase (MPnS) produces methylphosphonate, a metabolic precursor to methane in the upper ocean. Here, we determine a 2.35-angstrom resolution structure of MPnS and discover that it has an unusual 2-histidine-1-glutamine iron-coordinating triad.

O-H Activation by an Unexpected Ferryl Intermediate during Catalysis by 2-Hydroxyethylphosphonate Dioxygenase.

Activation of O-H bonds by inorganic metal-oxo complexes has been documented, but no cognate enzymatic process is known. Our mechanistic analysis of 2-hydroxyethylphosphonate dioxygenase (HEPD), which cleaves the C1-C2 bond of its substrate to afford hydroxymethylphosphonate on the biosynthetic pathway to the commercial herbicide phosphinothricin, uncovered an example of such an O-H-bond-cleavage event. Stopped-flow UV-visible absorption and freeze-quench Mössbauer experiments identified a transient iron(IV)-oxo (ferryl) complex.

Go it alone: four-electron oxidations by mononuclear non-heme iron enzymes.

This review discusses the current mechanistic understanding of a group of mononuclear non-heme iron-dependent enzymes that catalyze four-electron oxidation of their organic substrates without the use of any cofactors or cosubstrates. One set of enzymes acts on α-ketoacid-containing substrates, coupling decarboxylation to oxygen activation. This group includes 4-hydroxyphenylpyruvate dioxygenase, 4-hydroxymandelate synthase, and CloR involved in clorobiocin biosynthesis.

Oxygen-18 Kinetic Isotope Effects of Nonheme Iron Enzymes HEPD and MPnS Support Iron(III) Superoxide as the Hydrogen Abstraction Species.

Nonheme iron oxygenases that carry out four-electron oxidations of substrate have been proposed to employ iron(III) superoxide species to initiate this reaction [Paria, S.; Que, L.; Paine, T.

A common late-stage intermediate in catalysis by 2-hydroxyethyl-phosphonate dioxygenase and methylphosphonate synthase.

2-Hydroxyethylphosphonate dioxygenase (HEPD) and methylphosphonate synthase (MPnS) are nonheme iron oxygenases that both catalyze the carbon-carbon bond cleavage of 2-hydroxyethylphosphonate but generate different products. Substrate labeling experiments led to a mechanistic hypothesis in which the fate of a common intermediate determined product identity. We report here the generation of a bifunctional mutant of HEPD (E176H) that exhibits the activity of both HEPD and MPnS.

Structure and function of phosphonoacetaldehyde dehydrogenase: the missing link in phosphonoacetate formation.

Phosphonates (C-PO₃²⁻) have applications as antibiotics, herbicides, and detergents. In some environments, these molecules represent the predominant source of phosphorus, and several microbes have evolved dedicated enzymatic machineries for phosphonate degradation. For example, most common naturally occurring phosphonates can be catabolized to either phosphonoacetaldehyde or phosphonoacetate, which can then be hydrolyzed to generate inorganic phosphate and acetaldehyde or acetate, respectively.

Evidence that the fosfomycin-producing epoxidase, HppE, is a non-heme-iron peroxidase.

The iron-dependent epoxidase HppE converts (S)-2-hydroxypropyl-1-phosphonate (S-HPP) to the antibiotic fosfomycin [(1R,2S)-epoxypropylphosphonate] in an unusual 1,3-dehydrogenation of a secondary alcohol to an epoxide. HppE has been classified as an oxidase, with proposed mechanisms differing primarily in the identity of the O2-derived iron complex that abstracts hydrogen (H•) from C1 of S-HPP to initiate epoxide ring closure. We show here that the preferred cosubstrate is actually H2O2 and that HppE therefore almost certainly uses an iron(IV)-oxo complex as the H• abstractor.

Phosphonate biosynthesis and catabolism: a treasure trove of unusual enzymology.

Natural product biosynthesis has proven a fertile ground for the discovery of novel chemistry. Herein we review the progress made in elucidating the biosynthetic pathways of phosphonate and phosphinate natural products such as the antibacterial compounds dehydrophos and fosfomycin, the herbicidal phosphinothricin-containing peptides, and the antimalarial compound FR-900098. In each case, investigation of the pathway has yielded unusual, and often unprecedented, biochemistry.

Discovery and biosynthesis of phosphonate and phosphinate natural products.

The P-C bonds in phosphonate and phosphinate natural products endow them with a high level of stability and the ability to mimic phosphate esters and carboxylates. As such, they have a diverse range of enzyme targets that act on substrates containing such functionalities. Recent years have seen a renewed interest in discovery efforts focused on this class of compounds as well as in understanding their biosynthetic pathways.

Mechanistic investigation of methylphosphonate synthase, a non-heme iron-dependent oxygenase.

Methylphosphonate synthase is a non-heme iron-dependent oxygenase that converts 2-hydroxyethylphosphonate (2-HEP) to methylphosphonate. On the basis of experiments with two enantiomers of a substrate analog, 2-hydroxypropylphosphonate, catalysis is proposed to commence with stereospecific abstraction of the pro-S hydrogen on C2 of the substrate. Experiments with isotopologues of 2-HEP indicate stereospecific hydrogen transfer of the pro-R hydrogen at C2 of the substrate to the methyl group of methylphosphonate.

Mechanism and substrate recognition of 2-hydroxyethylphosphonate dioxygenase.

HEPD belongs to the superfamily of 2-His-1-carboxylate non-heme iron-dependent dioxygenases. It converts 2-hydroxyethylphosphonate (2-HEP) to hydroxymethylphosphonate (HMP) and formate. Previously postulated mechanisms for the reaction catalyzed by HEPD cannot explain its conversion of 1-HEP to acetylphosphate.

On the stereochemistry of 2-hydroxyethylphosphonate dioxygenase.

Stereochemical investigations have shown that the conversion of 2-hydroxyethylphosphonate to hydroxymethylphosphonate by the enzyme HEPD involves removal of the pro-S hydrogen at C2 and, surprisingly, the loss of stereochemical information at C1. As a result, the mechanisms previously proposed for HEPD must be re-evaluated.