Stream faunal assemblage structure is tied closely to hydrology and associated physiochemical properties. By altering natural flows, dams and their impoundments impact faunal assemblages over long distances. Although numerous studies have assessed the effect of dams on stream fauna, information is lacking for crayfishes. In this study, we characterized the effects of relatively large storage dams on crayfish assemblage structures. Over 2 years, we sampled three impounded and two unimpounded streams across two drainages in Alabama, United States, to identify biotic and abiotic factors correlated with crayfish assemblage metrics. Compared to impounded streams, unimpounded streams had greater habitat complexity (e.g., aquatic vegetation and woody debris), fewer predator fishes, lower minimum temperatures, and more variable discharges. These characteristics correlated with a higher density and diversity of crayfishes and smaller adults in impounded compared to unimpounded streams. Crayfish species assemblages differed between drainages, as did the biotic and abiotic factors affecting crayfish assemblages in each drainage, suggesting that these factors were species‐specific in their effects. Additionally, analysis of land uses suggested that factors other than dams may have also contributed to the observed differences in assemblage structures between impounded and unimpounded streams. For instance, in the more urbanized drainage, crayfish assemblages were more similar between up and downstream sections in all streams, regardless of impoundment. Our results indicate that large dams alter stream crayfish assemblage structure, with potentially cascading effects in trophic and organic matter dynamics both up and downstream.