The role of dynamical flexibility at the active site of a proteolytic enzyme alpha-chymotrypsin (CHT) has been correlated with its catalytic activity. The temperature-dependent efficiency of catalysis reveals a bell-shaped feature with a peak at 37 degrees C, the typical body temperature of homeothermal animals. The overall structural integrity of the enzyme in our experimental temperature range has been confirmed from dynamic light scattering (DLS) and circular dichroism (CD) studies. We have followed the dynamical evolution at the active site of CHT with temperature using picosecond-resolved fluorescence anisotropy of anthraniloyl probe (covalently attached to the serine-195 residue) and a substrate mimic (inhibitor) proflavin. The conformational dynamics at the active site is found to have a distinct connection with the enzyme functionality. The conformational flexibility of the enzyme is also evidenced from the compressibility studies on the enzyme. The site selective fluorescence detected circular dichroism (FDCD) studies reveal that the conformational flexibility of the enzyme has an effect on the structural perturbation at the active site. We have also proposed the possible implications of the dynamics in the associated energetics.