Dihydroneopterin aldolase (DHNA), an enzyme in the pathway that generates folic acid in bacteria, is investigated by a series of molecular dynamics simulations in its free form and complexed with its product, 6-hydroxymethyl-7,8-dihydropterin (HP). The active sites in DHNA are formed at the interface between pairs of protomers in this octameric protein. On the basis of root-mean-square deviation and root-mean-square fluctuation analyses of the trajectories, which take advantage of the presence of eight active sites, flexible regions of the apo protein surrounding the active site are identified and, upon binding HP, show that the active site is rigidified. Specific residues, associated with binding and the catalytic mechanism of DHNA, are associated with these flexible regions, and their interactions with HP account for most of the binding energy. A Principal Component Analysis shows rigidification of DHNA upon HP binding and that only a few modes of motion capture most of the atomic fluctuations in both apo and HP-bound forms. HP is pushed out of the active site in a series of simulations with different restrained positions between HP and DHNA to obtain a view of the exit pathway and energetic barrier to product release. The chosen pathway leads to a minimal disturbance of the system and provides a barrier consistent with the experimentally determined rate of product release. An analysis of the various components that contribute to the exit path energy and entropy provides insight into the energy-entropy compensation for product release.