Stability Engineering of Ferulic Acid Decarboxylase Unlocks Enhanced Aromatic Acid Decarboxylation.

George W. Roberts, Karl Fisher, Thomas Jowitt, David Leys

Research output: Contribution to journalArticlepeer-review

Abstract

Ferulic acid decarboxylase (Fdc) is a member of the microbial UbiD superfamily, a diverse family of (de)carboxylases capable of reversible decarboxylation on α,β-unsaturated acids. Recent application of Fdc includes in vivo generation of hydrocarbons such as isobutene and 1,3-butadiene, as well as C-H activation through CO² fixation. Protein engineering has expanded the substrate scope of the Aspergillus niger ferulic acid decarboxylase (AnFdc) to include (hetero)aromatic acid substrates. To further improve activity with aromatic acids, we introduced disulphide bonds into AnFdc to generate more thermostable variants. While some variants are negatively affected in co-factor incorporation and thus activity, others display increased thermostability and enhanced activity. The most thermostable disulphide bond AnFdc variant was combined with key active site mutations, allowing access to improved (hetero)aromatic decarboxylation including naphthoic acid decarboxylation. The reverse process, naphthalene carboxylation, is relevant to understanding microbial UbiD-mediated anaerobic naphthalene/benzene degradation. The improved naphthoic acid decarboxylation achieved here suggests further scope for AnFdc evolution towards an amenable model system for aromatic C-H activation through carboxylation.
Original languageEnglish
JournalCurrent Research in Chemical Biology
Publication statusAccepted/In press - 11 Aug 2023

Keywords

  • Fdc
  • prFMN
  • (de)carboxylase
  • thermostability
  • protein engineering
  • naphthoic acid decarboxylation

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