A bacterial flavin‐dependent oxidoreductase that captures carbon dioxide into biomass

Abstract

Atmospheric carbon dioxide is used as a carbon source for building biomass in plants and photosynthetic microbes. Non‐photosynthetic processes that also fix carbon dioxide have more recently been discovered. This research focuses on a microbial mechanism for coupling acetone to CO2 to make a central metabolite, acetoacetate. The key reaction is catalyzed by NADPH‐2‐ketopropyl‐coenzyme M oxidoreductase/carboxylase (2‐KPCC), a bacterial enzyme that is part of the flavin and cysteine‐disulfide containing oxidoreductase family (DSORs) which are best known for reducing metallic or disulfide substrates. Our research asks: how has nature repurposed a DSOR to, uniquely, break a C‐S bond and then trap and fix CO2? 2‐KPCC lacks a conserved, catalytically essential acid‐base histidine possessed by all other DSOR enzymes, having instead a phenylalanine (F501) at the same position. Mutagenesis showed that a F501H mutant has a similar rate of catalytic turnover; however, the product is acetone instead of acetoacetate (Figure 1). We hypothesized that F501 is important for both the reductive half reaction – which generates the reactive, C‐S bond breaking form of the active site – and for the oxidative half, in which an enolacetone intermediate reacts with either the correct (CO2) or incorrect (H+) electrophile. In this study, we used real‐time and spectroscopic methods to examine the reductive half reaction. In typical DSORs, this reaction generates a Cys/FAD charge transfer species. However, we showed that 2‐KPCC generates an electronically unique form of the active site, in which the flavin is oxidized and a pair of active site histidines are reduced and protonated. We hypothesize that this form of the active site generates the substrate‐reactive Cys in a more nucleophilic form, where it is capable of cleaving relatively strong C‐S bonds. The resulting enolacetone carbanion is an extremely potent nucleophile that is capable of directly attacking CO2. Research on 2‐KPCC and other biological CO2 fixation methods adds to our arsenal of strategies for carbon dioxide capture and use.Support or Funding InformationDOE:DE‐FG02‐04ER15563This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.