Enzymatic Reactions Involving Ketyls: From a Chemical Curiosity to a General Biochemical Mechanism

Abstract

Ketyls are radical anions with nucleophilic properties. Ketyls obtained by enzymatic one-electron reduction of thioesters were proposed as intermediates for the dehydration of (R)-2hydroxyacyl-CoA to (E)-2-enoyl-CoA. This concept was extended to the Birch-like reduction of benzoyl-CoA to 1,5-cyclohexadienecarboxyl-CoA. Nature uses two methods to achieve the therefore required low reduction potentials of less than −600 mV, either by an ATPdriven electron transfer similar to that catalyzed by the iron protein of nitrogenase or by electron bifurcation. Ketyls formed by thiyl radicalinitiated oxidation of alcohols followed by deprotonation are involved in coenzyme B 12 -independent diol dehydratases, other glycyl radical enzymes mediating key reactions in the degradations of choline, taurine, and 4-hydroxyproline, and all three classes of ribonucleotide reductases. A special case is the dehydration of 4hydroxybutyryl-CoA to crotonyl-CoA, which most likely proceeds via an oxidation to an allylic ketyl but requires neither a strong reductant nor an external radical generator. ■ KETYLS IN ORGANIC CHEMISTRYKetyls are radical anions, whichas the name impliesare derived from ketones by insertion of one electron into the antibonding 2p z orbital of the carbonyl group. 1 Two resonance structures contribute to the stability of a ketyl, the radical at carbon and the negative charge at oxygen and vice versa (Figure 1). Because the oxygen is more electronegative than

Diverse Energy-Conserving Pathways in Clostridium difficile: Growth in the Absence of Amino Acid Stickland Acceptors and the Role of the Wood-Ljungdahl Pathway

“…The reductive pathway relies on a "radicals by reduction" (86) strategy that involves ATP-dependent low-potential activation of the key enzyme 2-hydroxyisocaproyl-CoA dehydratase, whose catalytic role is to generate a remarkable ketyl radical intermediate from the substrate 2-hydroxyisocaproyl-CoA, which can then undergo dehydration and subsequent conversion to the product isocaprenoyl-CoA (87,88). Thereafter, reduction of isocaprenoyl-CoA and conversion to isocaproic acid proceeds by way of flavin-based electron bifurcation to regenerate 2 NAD ϩ and capture two low-potential equivalents in reduced ferredoxin (89,90). Growth in the absence of leucine deprives C. difficile of this very efficient reductive pathway and, at the same time, also requires a biosynthetic route to supply leucine for anabolic processes.…”

Elucidation of an anaerobic pathway for metabolism of L-carnitine-derived γ-butyrobetaine to trimethylamine in human gut bacteria

“…Yet there are notable distinctions in the substrate activation steps and key lyase enzymes of each pathway. Conversion of the carboxylate to the thioester lowers the pKa of the Cα-protons (25,26), facilitating elimination of the C4 hydroxyl or trimethylammonium group in the 4HBDor BbuA-catalyzed reaction, respectively. Due to its positive charge, the trimethylammonium moiety of γbb-CoA is a good leaving group for C4 elimination.…”

“…Based on the predicted annotations of the bbu genes, we hypothesized that γbb would be initially activated for subsequent TMA elimination through formation of a CoA thioester. The conversion of a carboxylic acid to a thioester is a common first step in many metabolic pathways as it lowers the pKa of the Cα-protons, facilitating further reactions (25,26). Indeed, in resting cell suspensions incubated with γbb, a metabolite was detected by LC-MS with a peak retention time and mass-to-charge ratio (895.5 m/z, [M+H] + ) matching those of a γbb-CoA standard ( Figure 3A, Figure S3).…”

Formation of Unstable and very Reactive Chemical Species Catalyzed by Metalloenzymes: A Mechanistic Overview

“…At the end of this reaction, a tyrosyl radical is generated, and the radical is transferred over 35 Å along the protein to the active site of the enzyme where it is finally accepted by a cysteine residue [156,161].…”