ABSTRACT
A three-step one-pot biocatalytic cascade was designed for the enantioselective formal α-amination of hexanoic acid to l-norleucine. Regioselective hydroxylation by P450CLA peroxygenase to 2-hydroxyhexanoic acid was followed by oxidation to the ketoacid by two stereocomplementary dehydrogenases. Combination with final stereoselective reductive amination by amino acid dehydrogenase furnished l-norleucine in >97% ee.
Subject(s)
Biocatalysis , Caproates/chemistry , Cytochrome P-450 Enzyme System/metabolism , Norleucine/chemistry , Amination , Bacteria/enzymology , Stereoisomerism , Substrate SpecificityABSTRACT
Selenoneine, a naturally occurring form of selenium, is the selenium analogue of ergothioneine, a sulfur species with health relevance not only as a purported antioxidant but likely also beyond. Selenoneine has been speculated to exhibit similar effects. To study selenoneine's health properties as well as its metabolic transformation, the pure compound is required. Chemical synthesis of selenoneine, however, is challenging and biosynthetic approaches have been sought. We herein report the biosynthesis and isolation of selenoneine from genetically modified fission yeast Schizosaccharomyces pombe grown in a medium containing sodium selenate. After cell lysis and extraction with methanol, selenoneine was purified by three consecutive preparative reversed-phase HPLC steps. The product obtained at the mg level was characterised by high resolution mass spectrometry, NMR and HPLC/ICPMS. Biosynthesis was found to be a promising alternative to chemical synthesis, and should be suitable for upscaling to produce higher amounts of this important selenium species in the future.
Subject(s)
Histidine/analogs & derivatives , Organoselenium Compounds/isolation & purification , Schizosaccharomyces/genetics , Schizosaccharomyces/metabolism , Chromatography, High Pressure Liquid , Genetic Engineering , Histidine/biosynthesis , Histidine/isolation & purification , Mass Spectrometry , Schizosaccharomyces/growth & developmentABSTRACT
A case of hydride-independent reaction catalyzed by flavin-dependent ene-reductases from the Old Yellow Enzyme (OYE) family was identified. α-Angelica lactone was isomerized to the conjugated ß-isomer in a nicotinamide-free and hydride-independent process. The catalytic cycle of C=C bond isomerization appears to be flavin-independent and to rely solely on a deprotonation-reprotonation sequence through acid-base catalysis. Key residues in the enzyme active site were mutated and provided insight on important mechanistic features. The isomerization of α-angelica lactone by OYE2 in aqueous buffer furnished 6.3â mm ß-isomer in 15â min at 30 °C. In presence of nicotinamide adenine dinucleotide (NADH), the latter could be further reduced to γ-valerolactone. This enzymatic tool was successfully applied on semi-preparative scale and constitutes a sustainable process for the valorization of platform chemicals from renewable resources.
Subject(s)
4-Butyrolactone/analogs & derivatives , Carbon/chemistry , Hydrogen/chemistry , Lactones/chemistry , Oxidoreductases/metabolism , 4-Butyrolactone/chemistry , Catalytic Domain , Isomerism , Oxidoreductases/chemistry , TemperatureABSTRACT
The asymmetric bioreduction of activated C-C-bonds catalyzed by a single flavoprotein was achieved via direct hydrogen transfer from a sacrificial 2-enone or 1,4-dione as hydrogen donor without requirement of a nicotinamide cofactor. Due to its simplicity, this system has clear advantages over conventional FAD-recycling systems.
