Cross species computational analysis of dopamine and MPP plus binding to the dopamine transporter elucidates mechanisms of MPTP induced neurotoxicity

1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces parkinsonism through its toxic metabolite, 1-methyl-4-phenylpyridinium (MPP+), which structurally mimics dopamine and enters dopaminergic neurons via the dopamine transporter (DAT). Although zebrafish are increasingly used as an MPTP-induced parkinsonian model due to their genetic and functional similarity to humans, direct molecular evidence supporting MPP+ transport via zebrafish DAT remains limited. This study computationally evaluated and compared the species-specific interactions of MPP+ and dopamine with human and zebrafish DAT using a multi-step in silico workflow comprising molecular docking, binding pocket identification, and molecular dynamics–based flexibility analysis. Docking was performed using AutoDock Vina, with human DAT (UniProt ID Q01959) and zebrafish DAT (UniProt ID Q90ZV1) prepared using AutoDock Tools. MPP+ (CID_39484) and dopamine (CID_681) were parameterized using the General AMBER Force Field (GAFF) tool. Blind docking employed identical 40 × 40 × 40 Å grids, and interactions were visualized using PyMol and LigPlot⁺. MPP+ exhibited binding affinities comparable to dopamine in both species (human: –7.2 vs –6.1 kcal/mol; zebrafish: –7.6 vs –6.4 kcal/mol) and showed substantial overlap in binding residues. Using CASTpFold, both ligands were found to occupy well-defined structural binding pockets that corresponded closely to docking-identified interaction residues, confirming biologically plausible binding cavities. To evaluate the dynamic stability of these interactions, coarse-grained flexibility analysis (CABS-flex) and normal mode analysis (iMODS) were conducted. RMSF profiles indicated that the DAT binding cavity remains intrinsically rigid, with ligand binding producing only minor local fluctuations in both species. iMODS further showed that dopamine- and MPP+-bound complexes shared similar deformability, B-factor distributions, variance patterns, and elastic network organization, with only slight differences in global stiffness between ligands. Collectively, these findings demonstrate that MPP+ closely mimics dopamine binding behaviour in both human and zebrafish DAT, supporting its ability to exploit DAT-mediated transport across species. Overall, this work provides integrated computational evidence—spanning docking, pocket geometry, and dynamic behaviour—that MPP+ engages DAT in a dopamine-like manner in both humans and zebrafish. This reinforces the validity of zebrafish as a model for studying dopaminergic neurotoxicity and Parkinson’s disease (PD). Nonetheless, the static and coarse-grained nature of the analyses highlights the need for future full all-atom molecular dynamics (MD) simulations and experimental validation to fully characterise MPP+ transport dynamics.