Molecular hydrogen (H₂) is an important metabolite cycled by microorganisms within the human gastrointestinal tract (GIT) with key roles in human nutrition and health. H2 is produced during fermentation by various bacteria and consumed as an energy source by other microbes, including acetogens, methanogens, and sulphate reducers. H₂ has traditionally been used as an indicator of gut dysbiosis through breath tests and disruption of H₂ cycling is associated with colorectal cancer, IBD and other GIT disorders. Despite links to human health, the microorganisms, pathways, and enzymes responsible for gastrointestinal H2 production remain unresolved. Here we show that a previously uncharacterised enzyme, the Group B [FeFe]-hydrogenases encoded by all four dominant gut phyla, primarily mediates fermentative H2 production in gut microbiota. Leveraging a dataset of 402 stool and biopsy metagenomes, 78 metatranscriptomes, and 812 gut bacterial isolate genomes, we show the genes encoding this enzyme are abundant, highly expressed, and widely distributed in the human GIT. Transcriptomic and gas chromatography analysis of 18 taxonomically diverse gut isolates confirms that the Group B [FeFe]- hydrogenase mediates rapid H2 production during fermentative growth. Furthermore, Bacteroides, a genus previously unknown to be hydrogenogenic, are dominant H₂ producers in this environment. Additional biochemical characterisation confirmed that the Group B [FeFe]-hydrogenase is catalytically active and binds a di-iron centre. This combination of culture-dependent and culture-independent analysis provides new insights into how H₂ is produced within the human GIT, and identifies key bacterial groups involved, enhancing our ever-growing understanding of the gut microbiota's impacts on human health.