Cofactor Generation Cascade for α-Ketoglutarate and Fe(II)-Dependent Dioxygenases

Florian Busch, Florian Busch, Jan Brummund, Elia Calderini, Martin Schürmann, Robert Kourist*

*Korrespondierende/r Autor/in für diese Arbeit

Publikation: Beitrag in einer FachzeitschriftArtikel

Abstract

Fe(II)- and α-ketoglutarate dependent dioxygenases have emerged as important catalysts for the preparation of non-natural amino acids. The stoichiometric supply of the cosubstrate α-ketoglutarate (αKG) is an important cost factor. A combination of the N-succinyl amino acid hydroxylase SadA with an l-glutamate oxidase (LGOX) allowed for coupling in situ production of αKG to stereoselective αKG-dependent dioxygenases in a one-pot/two-step cascade reaction. Both enzymes were used as immobilized enzymes and tested in a preparative scale setup under process-near conditions. Oxygen supply, enzyme, and substrate loading of the oxidation of glutamate were investigated under controlled reaction conditions on a small scale before upscaling to a 1 L stirred tank reactor. LGOX was applied with a substrate concentration of 73.6 g/L (339 mM) and reached a space-time yield of 14.2 g/L/h. Additionally, the enzyme was recycled up to 3 times. The hydroxylase SadA reached a space-time yield of 1.2 g/L/h at a product concentration of 9.3 g/L (40 mM). For both cascade reactions, the supply with oxygen was identified as a critical parameter. The results underline the robustness and suitability of α-ketoglutarate dependent dioxygenases for application outside of living cells.

Originalspracheenglisch
Seiten (von - bis)8604-8612
Seitenumfang9
FachzeitschriftACS Sustainable Chemistry and Engineering
Jahrgang8
Ausgabenummer23
DOIs
PublikationsstatusVeröffentlicht - 15 Jun 2020

ASJC Scopus subject areas

  • !!Chemistry(all)
  • Umweltchemie
  • !!Chemical Engineering(all)
  • !!Renewable Energy, Sustainability and the Environment

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