Strong evidence for the importance of scaffold conformational entropy in catalysis emerges from a series of time-resolved crystallographic, NMR, and computational studies of the homodimeric enzyme, fluoroacetate dehalogenase (FaDH) . Although the enzyme is dimeric, upon substrate binding at one active site, the other protomer does not bind substrate and can be thought of as an extended part of the scaffold. With clever mutations and substrate-analogues, these studies show how both solvent and protein scaffold entropy support the active site organization responsible for the chemical step of catalysis. This change to the scaffold occurs without any major backbone changes in any part of the catalytic cycle, but is evident in changes in: B-factors, alternative conformations of side chains, the number of bound waters, and NMR measurements of dynamics. Importantly, it is the contributions of distal residues and distal waters, not just waters displaced by the small molecules bound in the active site, that drive this entropic contribution. In addition, an analysis of MD trajectories using rigidity theory identified interprotomer allosteric pathways that suggests the mechanism of how these changes occur. Collectively these studies reveal how entropy can favor conformations responsible for rate-limiting steps of the catalytic cycle via increased conformational multiplicity of the scaffold and the release of solvent. The interplay between solvent and scaffold calls attention to the importance of holistically modeling the conformational dynamics of the entire system, not just the active site.