Clostridium sporogenes uses reductive Stickland metabolism in the gut to generate ATP and produce circulating metabolites

Abstract

Gut bacteria face a key problem in how they capture enough energy needed to sustain their growth and physiology. The gut bacterium Clostridium sporogenes obtains its energy by utilizing amino acids in pairs, coupling the oxidation of one to the reduction of another – the Stickland reaction. Oxidative pathways produce ATP via substrate level phosphorylation, whereas reductive pathways are thought to balance redox. Here, we investigated whether these reductive pathways are also linked to energy generation and the production of microbial metabolites that may circulate and impact host physiology. Using metabolomics, we find that during growth in vitro, C. sporogenes produces 15 metabolites, 13 of which are present in the gut of C. sporogenes colonized mice. Four of these compounds are reductive Stickland metabolites that circulate in the blood of gnotobiotic mice and are also detected in plasma from healthy humans. Gene clusters for reductive Stickland pathways suggest involvement of electron transfer proteins, and experiments in vitro demonstrate that reductive metabolism is coupled to ATP formation and not just redox balance. Genetic analysis points to the broadly conserved Rnf complex as a key coupling site for energy transduction. Rnf complex mutants show aberrant amino acid metabolism in defined medium and are attenuated for growth in the mouse gut, demonstrating a role of the Rnf complex in Stickland metabolism and gut colonization. Our findings reveal that the production of circulating metabolites by a commensal bacterium within the host gut is linked to an ATP-yielding redox process.

Eight Unexpected Selenoprotein Families in ABC transport, in Organometallic Biochemistry in Clostridium difficile and other anaerobes, and in Methylmercury Biosynthesis

“…Said organometallic compound might typically have a short half-life in solution, but is chaperoned before, during, and after transport by selenoproteins that have multiple Cys or Sec residues in contact with the metal atom. Following import, the compound is reduced by an 8Fe-9S (or possibly a 2(4Fe-4S)-Se) cluster-containing double-cubane protein, making the compound an oxidant that enables ATP generation by other cellular machinery, such as the energy-transducing Rnf complex (45). Once the compound can no longer be reduced further, the carbon-metal bond may be cleaved by SaoL, chaperoned to the efflux transporter SaoE, and extruded.…”

“…Following import, the compound is reduced by an 8Fe-9S (or possibly a 2(4Fe-4S)-Se) cluster-containing double-cubane protein, making the compound an oxidant that enables ATP generation by other cellular machinery, such as the energy-transducing Rnf complex (45). Once the compound can no longer be reduced further, the carbon-metal bond may be cleaved by SaoL, chaperoned to the efflux transporter SaoE, and extruded.…”

“…In Stickland fermentations, oxidation of one amino acid is coupled to reduction of another (or of sarcosine or betaine). The reductive metabolism leads to ATP formation (45). SAO operons are present in 0.3% of bacterial species, but in 13.4% of those with GrdX family proteins.…”

A widely distributed gene cluster compensates for uricase loss in hominids

“…Gibson assemblies were transformed into E. coli by electroporation, selected on LB-chloramphenicol (25 µg/mL) plates and sequenced to confirm the correct retargeted sequence. Intron re-targeted plasmids were transformed into E. coli s17-1 λpir and subsequently conjugated into C. sporogenes as described before (Liu et al, 2022). Mutants were verified by PCR and Sanger sequencing using the sequencing primers listed in Table S21 .…”

Systems biology illuminates alternative metabolic niches in the human gut microbiome

“…The growth strategies of individual gut species and strains shape their ability to colonize a host and their potential chemical interactions with other community members and with the host (Alexander et al, 2021; Medlock et al, 2018). Efforts to describe and model the metabolism and growth of various community members have included detailed biochemical studies of resource utilization by individual model species such as members of the genus Bacteroides (Koropatkin et al, 2012) and Clostridium sporogenes (Liu et al, 2022), as well as large-scale efforts to characterize species-level metabolic activity using community multi-omic profiling (Franzosa et al, 2018; Hertel et al, 2019). However, these efforts have been most fruitful for members of the microbiota that are found at high abundance and with prior knowledge of well-annotated metabolic pathways.…”