Comparative molecular dynamics reveal ligand-dependent structural and dynamic determinants of dipeptidyl peptidase IV recognition and inhibition

Dipeptidyl peptidase 4 (DPP4/CD26) is a clinically relevant serine protease involved in metabolic, immunological and viral processes. Although several crystal structures of DPP4-ligand complexes have been reported, the dynamic features that distinguish substrates from inhibitors remain less understood. In this study, 250 ns all-atom molecular dynamics simulations were performed for four structurally distinct ligand-bound states, a synthetic construct, a Neuropeptide Y (NPY) fragment, Diprotin A and an HIV-1 TAT peptide, to examine how ligand identity affects the conformational behavior of DPP4. All complexes maintained the global α/β-hydrolase and β-propeller architecture, with secondary-structure content remaining stable throughout the simulations. Ligand-dependent differences were mainly observed in the flexible loop regions near the catalytic cavity. The inhibitory ligands (Diprotin A and TAT) promoted reduced mobility and lower backbone RMSD values (1.5-2.0 Å), whereas the substrate and the synthetic construct allowed greater local flexibility (2.0-2.5 Å). Contact analyses showed a conserved anchoring network involving Glu205, Glu206 and Tyr662 across all systems, along with ligand-specific peripheral interactions, including Trp659, Tyr752 and Phe357, that influence the binding modes. These observations reveal a relationship among structure-dynamics-function, in which DPP4 maintains a stable interaction and modulates loop movements, including peripheral contacts, to differentiate substrates from inhibitors. The dynamic behavior of the enzyme offers mechanistic insights that allow for specific ligand recognition, which could support the design and development of new DPP4 modulators.