NonhemeFe(II)‐ and 2‐Oxoglutarate‐Dependent Ethylene‐Forming Enzyme

Abstract

The ethylene‐forming enzyme (EFE) is a ferrous ion‐ and l ‐arginine ( l ‐Arg)‐dependent oxygenase that cleaves three carbon–carbon bonds of 2‐oxoglutarate (2OG) as it forms ethylene plus three molecules of carbon dioxide/bicarbonate. The enzyme also catalyzes the oxidative decarboxylation of 2OG (producing carbon dioxide and succinate) while hydroxylating the C5 position of l ‐Arg, with this intermediate species decomposing spontaneously to guanidine and L‐Δ‐1‐pyrroline‐5‐carboxylate. This secondary reaction is typical of the hydroxylation reactions catalyzed by a broad range of Fe(II)/2OG‐dependent oxygenases, whereas the ethylene‐generating reaction is unprecedented. Genes encoding homologs of EFE are present in specific fungi and bacteria, and the protein is proposed to be a virulence factor in some plant pathogens. The EFE sequence and structure are clearly related to those of 1‐aminocyclopropane‐1‐carboxylate oxygenase, an Fe(II)‐ and dioxygen‐dependent ethylene‐forming enzyme that does not require or utilize 2OG. By contrast, the EFE sequence is not significantly related to three Fe(II)/2OG‐dependent oxygenases (VioC, NapI, and OrfP) that catalyze other reactions with l ‐Arg (hydroxylation at C3, C4‐C5 desaturation, and C3,C4 dihydroxylation, respectively). The best characterized EFE is that from Pseudomonas syringae pv. phaseolicola strain PK2. A visible absorption spectrum associated with the Fe(II)/2OG metal‐to‐ligand charge‐transfer transition of EFE·Fe(II)·2OG is less intense than for other members of this enzyme family; this result is explained by the predominantly monodentate metal coordination of 2OG as seen in the corresponding EFE·Mn(II)·2OG crystal structure. Binding of l ‐Arg leads to an intensification of the Fe(II)/2OG visible absorption spectrum and a shift to bidentate metal coordination. In EFE·Mn(II)·2OG, the metal is bound by one oxygen atom of the Asp191 carboxylate, but in EFE·Mn(II)·2OG,· l ‐Arg this atom shifts to interact with l ‐Arg and the other carboxylate oxygen atom binds the metal. A reorientation of Tyr192 accompanies l ‐Arg binding, forming a hydrogen bond between the phenol hydroxyl group and the amino acid. The carboxylate shift and the aromatic side chain reorientation lead to a twisted Asp191‐Tyr192 peptide bond. The environment of 2OG in EFE is more hydrophobic compared to other family members. In combination, the altered position of the metal‐binding carboxylate, the twisted peptide bond, and the hydrophobic environment surrounding 2OG may all contribute to the ability of EFE to catalyze ethylene formation.

Biological formation of ethylene

“…1B). 11,12 Its primary reaction is to convert 2-oxoglutarate (2OG) to ethylene and three molecules of carbon dioxide/bicarbonate (CO 2 ) in an oxidative reaction that requires, but does not transform, the amino acid l -Arg. The secondary reaction catalyzed by EFE is to hydroxylate C5 of l -Arg, creating an unstable species that decomposes to guanidine and l -Δ 1 -pyrroline-5-carboxylic acid (P5C), while converting 2OG to succinate and carbon dioxide.…”