Pesci, LorenzoLorenzoPesciGurikov, PavelPavelGurikovLiese, AndreasAndreasLieseKara, SelinSelinKara2019-08-222019-08-222017-09-01Biotechnology Journal 12 (12): 1700332 (2017-12-01)http://hdl.handle.net/11420/3160A variety of strategies is applied to alleviate thermodynamic and kinetic limitations in biocatalytic carboxylation of metabolites in vivo. A key feature to consider in enzymatic carboxylations is the nature of the cosubstrate: CO₂ or its hydrated form, bicarbonate. The substrate binding and activation mechanism determine what the actual carboxylation agent is. Dihydroxybenzoic acid (de)carboxylases catalyze the reversible regio-selective ortho-(de)carboxylation of phenolics. These enzymes have attracted considerable attention in the last 10 years due to their potential in substituting harsh conditions typical of chemical carboxylations (100–200 °C, 5–100 bar) with, ideally, greener ones (20–40 °C, 1 bar). They are reported to use bicarbonate as substrate, needed in large excess to overcome thermodynamic and kinetic limitations. Therefore, CO₂ can be used as substrate by these enzymes only if it is converted into bicarbonate in situ. In this contribution, we report the simultaneous amine-mediated conversion of CO₂ into bicarbonate and the ortho-carboxylation of different phenolic molecules catalyzed by 2,3-dihydroxybenzoic acid (de)carboxylase from Aspergillus oryzae. Our results show that under the newly developed conditions a significant thermodynamic (up to twofold increase in conversion) and kinetic improvement (up to approx. fivefold increase in rate) of the biocatalytic carboxylation of catechol is achieved.en1860-6768201712Wiley-VCH2-hydroxybenzoic acidsacids amine scrubbingcarboxylation(de)carboxylasesenzyme kineticsBiowissenschaften, BiologieIngenieurwissenschaftenJournal Article10.1002/biot.201700332Other