Bacteria in reservoir systems are controlled by a variety of factors. In this study, a lacustrine system with high primary production fed by a river with lower algal biomass was used to examine the relationship between bacterial community composition (BCC) and function under varying environmental conditions. Bacterial abundance, respiration, production, and growth efficiency were measured in the Cuyahoga River and Lake Rockwell reservoir in Ohio (USA). BCC was determined via terminal restriction length polymorphisms of 16S rRNA genes. Also, in laboratory experiments, bacterial communities were exposed to algal exudates and leaf leachates. Concurrently with physicochemical differences between lacustrine and riverine sites, there were differences in BCC that seemed unrelated to environmental conditions and DOC sources (color: chlorophyll a ratio). Functional attributes varied among dates but not sites, except for bacterial heterotrophic productivity. Overall, the source of DOC differed between the river and reservoir as did BCC. In contrast, bacterial functions, with the exception of productivity, were similar between the river and reservoir sites. Environmental variables (e.g., temperature, nutrient, and chlorophyll a) were correlated with differences in bacterial function. Differences in BCC did not coincide with functional differences in the field or controlled laboratory conditions suggesting that structure and function were uncoupled.