Exploring the directionality of Escherichia coli formate hydrogenlyase: a membrane‐bound enzyme capable of fixing carbon dioxide to organic acid. Issue 5 (2nd May 2016)

• Record Type: Journal Article
• Authors: Pinske, Constanze; Sargent, Frank
• Is Part Of: MicrobiologyOpen. Volume 5:Issue 5(2016:Oct.)
• Page Start: 721
• Page End: 737
• Publication Date: 2016-05-02
• Subjects: [NiFe] hydrogenase -- bacterial hydrogen metabolism -- formate chemosynthesis -- formate dehydrogenase -- formate hydrogenlyase -- site‐directed mutagenesis; Microbiology -- Periodicals; 579
• DOI: 10.1002/mbo3.365
• Languages: English
• ISSNs: 2045-8827

Abstract

Abstract: During mixed‐acid fermentation Escherichia coli produces formate, which is initially excreted out the cell. Accumulation of formate, and dropping extracellular pH, leads to biosynthesis of the formate hydrogenlyase (FHL) complex. FHL consists of membrane and soluble domains anchored within the inner membrane. The soluble domain comprises a [NiFe] hydrogenase and a formate dehydrogenase that link formate oxidation directly to proton reduction with the release of CO2 and H2 . Thus, the function of FHL is to oxidize excess formate at low pH. FHL subunits share identity with subunits of the respiratory Complex I. In particular, the FHL membrane domain contains subunits (HycC and HycD) that are homologs of NuoL/M/N and NuoH, respectively, which have been implicated in proton translocation. In this work, strain engineering and new assays demonstrate unequivocally the nonphysiological reverse activity of FHL in vivo and in vitro. Harnessing FHL to reduce CO2 to formate is biotechnologically important. Moreover, assays for both possible FHL reactions provide opportunities to explore the bioenergetics using biochemical and genetic approaches. Comprehensive mutagenesis of hycC did not identify any single amino acid residues essential for FHL operation. However, the HycD E199, E201, and E203 residues were found to be critically important for FHL function. Abstract : The formate hydrogenlyase (FHL) complex of Escherichia coli has been characterized in vivo and in vitro. As well as performing hydrogen evolution linked to formate oxidation, FHL has now been shown to be capable of converting CO2 into formate using molecular hydrogen as reductant. The enzyme activity was analyzed in combination with a comprehensive mutagenic analysis of the subunits of the FHL membrane domain, HycC and HycD. FHL is probably not an energy‐conserving enzyme.