Engineering the promiscuous racemase activity of an arylmalonate decarboxylase

Robert Kourist, Yusuke Miyauchi, Daisuke Uemura, Kenji Miyamoto

Research output: Contribution to journalArticlepeer-review


Variant G74C of arylmalonate decarboxylase (AMDase) from Bordatella bronchoseptica has a unique racemising activity towards profens. By protein engineering, variant G74C/V43A with a 20-fold shift towards promiscuous racemisation was obtained, based on a reduced activity in the decarboxylation reaction and a two-fold increase in the racemisation activity. The mutant showed an extended substrate range, with a 30-fold increase in the reaction rate towards ketoprofen. Molecular dynamics simulations and the substrate profile of the racemase indicate that the steric and polar effects of the substrate structure play a more dominant role on catalysis than mere kinetic α-proton acidity. The observation that the conversion of β,γ-unsaturated carboxylic acids does not lead to a rearrangement to form their α,β isomers indicates a concerted rather than a stepwise mechanism. Interestingly, a substrate bearing a nitro group instead of the carboxylic acid group on the α-carbon atom was also converted by the racemase.

Original languageEnglish
Pages (from-to)557-63
Number of pages7
JournalChemistry - a European Journal
Issue number2
Publication statusPublished - 10 Jan 2011
Externally publishedYes


  • Bordetella bronchiseptica
  • Butyrates
  • Carboxy-Lyases
  • Catalysis
  • Computer Simulation
  • Genetic Variation
  • Models, Molecular
  • Molecular Structure
  • Mutagenesis, Site-Directed
  • Protein Engineering
  • Racemases and Epimerases
  • Stereoisomerism
  • Substrate Specificity
  • Journal Article
  • Research Support, Non-U.S. Gov't

Fields of Expertise

  • Human- & Biotechnology


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