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

Cross species computational analysis of dopamine and MPP plus binding to the dopamine transporter elucidates mechanisms of MPTP induced neurotoxicity | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Cross species computational analysis of dopamine and MPP plus binding to the dopamine transporter elucidates mechanisms of MPTP induced neurotoxicity Khairiah Razali, Mohd Hamzah Mohd Nasir, Jaya Kumar, Wael Mohamed This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8417718/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract 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. Full Text Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8417718","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":569448702,"identity":"f1163e5a-859f-4147-a696-5271f64297d2","order_by":0,"name":"Khairiah Razali","email":"","orcid":"","institution":"International Islamic University Malaysia","correspondingAuthor":false,"prefix":"","firstName":"Khairiah","middleName":"","lastName":"Razali","suffix":""},{"id":569448703,"identity":"2e6b1285-e972-45e8-bcc4-e41648fde161","order_by":1,"name":"Mohd Hamzah Mohd Nasir","email":"","orcid":"","institution":"International Islamic University Malaysia","correspondingAuthor":false,"prefix":"","firstName":"Mohd","middleName":"Hamzah Mohd","lastName":"Nasir","suffix":""},{"id":569448704,"identity":"ef66abf8-8a98-4da8-bf80-04c73336ab95","order_by":2,"name":"Jaya Kumar","email":"","orcid":"","institution":"National University of Malaysia","correspondingAuthor":false,"prefix":"","firstName":"Jaya","middleName":"","lastName":"Kumar","suffix":""},{"id":569448705,"identity":"066947ae-ef86-4448-b80a-b5a23c60af9b","order_by":3,"name":"Wael Mohamed","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAu0lEQVRIiWNgGAWjYJCCA4n/bIAUY+MBYnUwHvjAlgaiG4jWwnxwBtthiHVEqTe4dsbgMA/Pebu17YeBttTYRBPWcjsHqEXidvK2M4lALcfSchsIaTEDazG4nWx2AKiFseEwsVoSziWbnX9IgpaDMw4csDO7Qawt9rfTCg58bEhOMLsBtCWBGL9Izk7e/CGxwc7e7Hz6wwcfamwIa2Fg4DAAkYlglQmElYMA+wOwA4lTPApGwSgYBSMSAACP5k3ccN9G3QAAAABJRU5ErkJggg==","orcid":"","institution":"International Islamic University Malaysia","correspondingAuthor":true,"prefix":"","firstName":"Wael","middleName":"","lastName":"Mohamed","suffix":""}],"badges":[],"createdAt":"2025-12-21 14:08:28","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8417718/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8417718/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":99798544,"identity":"706c2e3d-563d-41c7-b418-9c90010c84d3","added_by":"auto","created_at":"2026-01-08 13:48:36","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1960405,"visible":true,"origin":"","legend":"","description":"","filename":"ManuscriptAmended.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8417718/v1_covered_384ede02-4b88-4547-ac6b-f7bf7f89e267.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Cross species computational analysis of dopamine and MPP plus binding to the dopamine transporter elucidates mechanisms of MPTP induced neurotoxicity","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-8417718/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8417718/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"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). 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