Biology subjects

Kountz, D. J.

Publications and source records attributed to Kountz, D. J..

4 recordsLinked to original sources

bioRxiv2025

Flavoaffinins, elusive cellulose-binding natural products from an anaerobic bacterium

Cellulose is the most abundant polymer on earth. Many anaerobic cellulose-degrading bacteria produce uncharacterized yellow-orange, cellulose-binding pigments known as yellow affinity substances (here referred to as flavoaffinins) that are associated with cellulose degradation. Here, we isolate and structurally characterize the flavoaffinins from Clostridium (Hungateiclostridium) thermocellum, a key workhorse for the industrial conversion of cellulosic feedstocks to ethanol. Flavoaffinins represent an unprecedented structural juxtaposition of an aryl polyene chain with a hydroxy-diene {gamma}-lactone. We also shed light on their biosynthesis using stable-isotope feeding experiments. This effort lays the groundwork for understanding the biological function(s) of the flavoaffinins and expands the limited number of natural products isolated from obligately anaerobic microbes.

biochemistry↗

bioRxiv2024

A widespread hydrogenase drives fermentative growth of gut bacteria in healthy people

Molecular hydrogen (H2) is among the most central, but least understood, metabolites in the human gastrointestinal tract (gut). H2 gas is produced in large quantities during bacterial fermentation and consumed as an energy source by bacteria and archaea. Disruption of H2 cycling is linked to gastrointestinal disorders, infections, and cancers, with H2 used as an indicator of gut dysfunction through breath tests. Despite this, the microorganisms, pathways, and enzymes mediating H2 production remain unresolved. Here we show that a previously uncharacterised enzyme, the group B [FeFe]-hydrogenase, drives most fermentative H2 production in the human gut. Analysis of stool, biopsy, and isolate (meta)genomes and (meta)transcriptomes show this hydrogenase is encoded by most gut bacteria and is highly expressed. Through analysis of 19 taxonomically diverse gut isolates, the group B [FeFe]-hydrogenase produces large amounts of H2 gas and supports fermentative growth of both Bacteroidetes and Firmicutes. Bacteroides particularly dominate H2 production. Biochemical and spectroscopic characterisation shows purified group B [FeFe]-hydrogenases are catalytically active and bind a di-iron active site. These hydrogenases are highly enriched in the guts of healthy individuals, but significantly depleted in favour of other fermentative hydrogenases in Crohns disease. Furthermore, we show that metabolically flexible respiratory bacteria are the most abundant H2 oxidizers in the gut, not sulfate reducers, methanogens, and acetogens as previously thought. This combination of enzymatic, cellular, and ecosystem-level analysis provides the first detailed understanding of H2 cycling in the human gut and reveals new links between microbiota function and gastrointestinal health.

microbiology↗

bioRxiv2023

A diversified, widespread microbial gene cluster encodes homologs of methyltransferases involved in methanogenesis.

Analyses of microbial genomes have revealed unexpectedly wide distributions of enzymes from specialized metabolism, including methanogenesis, providing exciting opportunities for discovery. Here, we identify a family of gene clusters (the type 1 mlp gene clusters (MGCs)) that encodes homologs of the soluble coenzyme M methyltransferases (SCMTs) involved in methylotrophic methanogenesis and is widespread in bacteria and archaea. Type 1 MGCs are expressed and regulated in medically, environmentally, and industrially important organisms, making them likely to be physiologically relevant. Enzyme annotation, analysis of genomic context, and biochemical experiments suggests these gene clusters play a role in methyl-sulfur and/or methyl-selenide metabolism in numerous anoxic environments, including the human gut microbiome, potentially impacting sulfur and selenium cycling in diverse, anoxic environments.

bioinformatics↗

bioRxiv2021

The Stickland fermentation precursor trans-4-hydroxyproline differentially impacts the metabolism of Clostridioides difficile and commensal Clostridia

An intact gut microbiota confers colonization resistance against Clostridioides difficile through a variety of mechanisms, likely including competition for nutrients. Recently, proline was identified as an important environmental amino acid that C. difficile uses to support growth and cause significant disease. A post-translationally modified form, trans-4-hydroxyproline, is highly abundant in collagen, which is degraded by host proteases in response to C. difficile toxin activity. The ability to dehydrate trans-4-hydroxyproline via the HypD glycyl radical enzyme is wide-spread amongst gut microbiota, including C. difficile and members of the commensal Clostridia, suggesting that this amino acid is an important nutrient in the host environment. Therefore, we constructed a C. difficile {Delta}hypD mutant and found that it was modestly impaired in fitness in a mouse model of infection, and was associated with an altered microbiota when compared to mice challenged with the wild type strain. Changes in the microbiota between the two groups were largely driven by members of the Lachnospiraceae family and the Clostridium genus. We found that C. difficile and type strains of three commensal Clostridia had significant alterations to their metabolic gene expression in the presence of trans-4-hydroxyproline in vitro. The proline reductase (prd) genes were elevated in C. difficile, consistent with the hypothesis that trans-4-hydroxyproline is used by C. difficile to supply proline for fermentation. Similar transcripts were also elevated in some commensal Clostridia tested, although each strain responded differently. This suggests that the uptake and utilization of other nutrients by the commensal Clostridia may be affected by trans-4-hydroxyproline metabolism, highlighting how a common nutrient may be a signal to each organism to adapt to a unique niche. Further elucidation of the differences between them in the presence of hydroxyproline and other key nutrients will be important to determining their role in nutrient competition against C. difficile.

microbiology↗