Drugs targeting transcriptional cofactors (TcoFs) function well in mouse models but fail to replicate their efficacy in human beings. Thus, we performed a comparative study on the TcoFs of humans and mice to find the similarity and dissimilarity between them. We observed high similarity in protein sequence and interacting domains between humans and mice. At the same time, dissimilarity was gradually increased in terms of interacting motifs, post-translational modifications, and molecular switches. Indeed, some of the post-translational modifications and molecular switches present in human beings are preferentially exempted in mice. Thus, structure-specific drugs designed to target TcoFs are functional in mice but fail in human beings, because the absence of some molecular switches in mice offers a particular conformation on the interacting motifs, which might facilitate drug binding. But in humans, owing to the presence of molecular switches, drug binding is not possible. From molecular dynamics simulation analysis, we inferred 8 different molecular switches on 3 proteins and found that 5 molecular switches influenced structural change in interacting motifs and revealed the reason for the functioning of drug in mice but not in human beings. From protein interaction network analysis, we find that a few interacting partners in mice are exempted in humans, and in both the cases the interacting partners are high when the domains are highly structured and the interacting partners are low when the domains are highly disordered. Hence, we are sure that our investigations will provide a promising support in future for designing drugs with high translational efficiency from mice models to human clinical trials. © 2018 Wiley Periodicals, Inc.