Recent studies of the bacterial enzymes EcMTAN and VcMTAN showed that they have different binding affinities for the same transition state analogue. This was surprising given the similarity of their active sites. We performed transition path sampling simulations of both enzymes to reveal the atomic details of the catalytic chemical step, which may be the key for explaining the inhibitor affinity differences. Even though all experimental data would suggest the two enzymes are almost identical, subtle dynamic differences manifest in differences of reaction coordinate, transition state structure, and eventually significant differences in inhibitor binding. Unlike EcMTAN, VcMTAN has multiple distinct transition states, which is an indication that multiple sets of coordinated protein motions can reach a transition state. Reaction coordinate information is only accessible from transition path sampling approaches, since all experimental approaches report averages. Detailed knowledge could have a significant impact on pharmaceutical design.