Aims: In a biofilm reactor, detachment force resulting from hydraulic shear is a major factor that determines the formation and structure of steady state biofilm. The metabolic response of biofilm to change in shear stress was therefore investigated. Methods and Results: A conventional annular reactor made of PVC was used, in which shearing over the rotating disc surface was strictly defined. Results from the steady state aerobic biofilm reactor showed that the biofilm structure (density and thickness) and metabolic behaviour (growth yield and dehydrogenase activity) were closely related to the shear stress exerted on the biofilm. Smooth, dense and stable biofilm formed at relatively high shear stress. Higher dehydrogenase activity and lower growth yield were obtained when the shear stress was raised. Growth yield was inversely correlated with the catabolic activity of biofilm. The reduced growth yield, together with the enhanced catabolic activity, suggests that a dissociation of catabolism from anabolism would occur at high shear stress. Conclusions: Biofilms may respond to shear stress by regulating metabolic pathways associated with the substrate flux flowing between catabolism and anabolism. A biological phenomenon, besides a simple physical effect, is underlying the observed relation between the shear stress and resulting biofilm structure. Significance and Impact of the Study: A hypothesis is proposed that the shear-induced energy spilling would be associated with a stimulated proton translocation across the cell membrane, which favours formation of a stronger biofilm. This research may provide a basis for experimental data on biofilm obtained at different shear stresses to be interpreted in relation to energy.