We explored the seasonal and vertical patterns of cell numbers, biomass, and amino acid incorporation of three major phylogenetic groups of the bacterioplankton in an oligomesotrophic alpine lake. Throughout the year the numerically dominant bacteria in the oxygenated epilimnion were small Actinobacteria. The timing and vertical location of their spring peak coincided with that of the heterotrophic flagellates (HNF). A second maximum of the relative proportions of Actinobacteria occurred during an autumn bloom of the diatom Asterionella formosa. Betaproteobacteria were negatively related to HNF abundances and formed maximal numbers and biomass during the summer stratification in the increasingly oxygen-depleted hypolimnion. Members of the Cytophaga-Flavobacteria subclade (CF) of Bacteroidetes preferably inhabited the completely anoxic zone of the hypolimnion. The numerically dominant Actinobacteria contributed only little to total bacterial biomass, whereas Betaproteobacteria and the comparatively rare (albeit large) CF represented the bulk of the bacterial carbon pool. Amino acid incorporation was mainly observed in Actinobacteria and Betaproteobacteria. Actinobacteria showed a clear preference for amino acids other than leucine. These bacteria represented the most active biomass (with respect to amino acid uptake), whereas the opposite was the case for CF. The contrasting vertical and seasonal patterns suggest that the three phylogenetic groups to some extent represent different functional compartments of the bacterioplankton.