Microbial communities play an essential role in marine biogeochemical cycles. Physical and biogeochemical changes in Jinhae Bay, the most anthropogenically eutrophied bay on the coasts of South Korea, are well described, but less is known about the associated changes in microbial communities. Temporal and vertical variation in microbial communities at three depths (surface, middle, and bottom) at seven time points (June to December) at the J1 sampling site were investigated on the MiSeq platform based on the 16S rRNA gene. Overall, the microbial community was dominated by Proteobacteria, Cyanobacteria, and Bacteroidetes from June to November, whereas Firmicutes were dominant in December, especially in the middle and bottom layers. The results indicate that the microbial community composition strongly varied with temporal changes in the physicochemical water properties. Moreover, the community composition differed markedly between the surface and middle layers and the bottom layer in the summer, when the water column was strongly stratified and bottom water hypoxia developed. A redundancy analysis suggested a significant correlation between physicochemical variables (i.e., temperature, salinity, and oxygen concentration) and microbial community composition. This study indicates that temporal changes in water conditions and eutrophication-induced hypoxia effectively shape the structure of the microbial community.