Hydrogen gas (H₂) is naturally produced by biological and non-biological reactions in various environmental niches. However, the influence of H₂ on microbial processes that cause the mobilization and release of arsenic from solid phase into groundwater remains to be resolved. Given that dissimilatory arsenate [As(V)]-respiring prokaryotes (DARPs) have been demonstrated to significantly contribute to the formation of As-contaminated groundwater, our study specifically examined the interactions between H₂ and DARPs. We prepared an enriched DARP population from As-contaminated soils. Metagenomic analyses of the DARP population revealed that approximately 46.7 % of the qualified DARPs’ MAGs contain at least one type I Ni-Fe hydrogenase. The Ni-Fe hydrogenase proteins in DARPs show unique diversity. Functional assays indicate that the DARP population exhibited notable activity in oxidizing H₂ while concurrently reducing As(V) under strictly anaerobic conditions. Arsenic release assays indicate that the DARP population is highly proficient at catalyzing the reductive mobilization of arsenic in scorodite, using hydrogen as the electron donor. These findings offer the initial evidence that H₂ can directly promote the formation of arsenic-contaminated groundwater mediated by DARPs, a biogeochemical process that has long been overlooked. Therefore, this study increases our insight into the microbial mechanisms involved in the formation of arsenic-contaminated groundwater.