2017
Conformational dynamics of a neurotransmitter:sodium symporter in a lipid bilayer
Adhikary S, Deredge DJ, Nagarajan A, Forrest LR, Wintrode PL, Singh SK. Conformational dynamics of a neurotransmitter:sodium symporter in a lipid bilayer. Proceedings Of The National Academy Of Sciences Of The United States Of America 2017, 114: e1786-e1795. PMID: 28223522, PMCID: PMC5347597, DOI: 10.1073/pnas.1613293114.Peer-Reviewed Original ResearchConceptsSodium symportersMembrane proteinsMammalian membrane proteinsConformational mechanismIntegral membrane proteinsPhospholipid bilayer nanodiscsSolution spectroscopyDetergent-solubilized stateExtracellular loop 2Site-specific labelingSmall molecule neurotransmittersBilayer nanodiscsHydrogen-deuterium exchangeX-ray crystallographyCysteine accessibilityConformational dynamicsMembrane mimicsMolecular dynamics simulationsLoop 2Endogenous cysteineHelices 1ALeuTLipid bilayersProteinSymporter
2015
Chapter Nine Biophysical Approaches to the Study of LeuT, a Prokaryotic Homolog of Neurotransmitter Sodium Symporters
Singh SK, Pal A. Chapter Nine Biophysical Approaches to the Study of LeuT, a Prokaryotic Homolog of Neurotransmitter Sodium Symporters. Methods In Enzymology 2015, 557: 167-198. PMID: 25950965, PMCID: PMC4818570, DOI: 10.1016/bs.mie.2015.01.002.Peer-Reviewed Original ResearchConceptsMultiple integral membrane proteinsSolute carrier 6 (SLC6) familyNeurotransmitter sodium symportersIntegral membrane proteinsAmino acid symporterDependent neurotransmitter transportersEukaryotic counterpartsSodium symportersProkaryotic homologAcid symporterLeuT structureMembrane proteinsNeurotransmitter transportersBiophysical approachesLeuTHomology modelingMechanism of transportSecondary transportAmino acidsLipid bilayersSignificant transportersMechanistic paradigmMolecular dynamics simulationsSymporterProtein
2013
Radioligand Binding to Nanodisc-Reconstituted Membrane Transporters Assessed by the Scintillation Proximity Assay
Nasr ML, Singh SK. Radioligand Binding to Nanodisc-Reconstituted Membrane Transporters Assessed by the Scintillation Proximity Assay. Biochemistry 2013, 53: 4-6. PMID: 24344975, PMCID: PMC4062192, DOI: 10.1021/bi401412e.Peer-Reviewed Original ResearchConceptsMembrane transportersPhospholipid bilayer nanodiscsNative cell membranesSecondary transportersScintillation proximityBilayer nanodiscsDetergent solubilizationCell membraneLipid vesiclesTransportersNanodiscsScreening effortsBackground interferenceLeuTProteinVesiclesPowerful techniqueBindingMembraneReconstitutionRadioligand bindingSolubilizationInterferencePotential challengesProximity
2008
A Competitive Inhibitor Traps LeuT in an Open-to-Out Conformation
Singh SK, Piscitelli CL, Yamashita A, Gouaux E. A Competitive Inhibitor Traps LeuT in an Open-to-Out Conformation. Science 2008, 322: 1655-1661. PMID: 19074341, PMCID: PMC2832577, DOI: 10.1126/science.1166777.Peer-Reviewed Original ResearchMeSH KeywordsAmino Acid Transport SystemsAmino AcidsBacterial ProteinsBinding SitesBinding, CompetitiveBiological TransportCrystallizationCrystallography, X-RayHydrogen BondingHydrophobic and Hydrophilic InteractionsKineticsLeucineLigandsModels, BiologicalModels, MolecularProtein ConformationProtein Structure, TertiarySodiumSymportersTryptophanConceptsNeurotransmitter sodium symportersPrimary substrate siteAmino acid substratesSodium symportersSecondary transportersExtracellular gateSubstrate passageLeucine transporterArginine 30Substrate transportCellular membranesConformational changesAcid substratesConformational statesSubstrate siteFunctional studiesIon gradientsLeuTWeak binding sitesTransportersBinding sitesSmall moleculesCompetitive inhibitorConformationTrp complexLeuT: A prokaryotic stepping stone on the way to a eukaryotic neurotransmitter transporter structure
Singh SK. LeuT: A prokaryotic stepping stone on the way to a eukaryotic neurotransmitter transporter structure. Channels 2008, 2: 380-389. PMID: 19066470, DOI: 10.4161/chan.2.5.6904.Peer-Reviewed Original ResearchConceptsSodium binding siteSolute carrier 6 (SLC6) familyIntegral membrane proteinsStructure/function studiesBinding sitesCrystal structureEukaryotic counterpartsAccurate homology modelsTransporter structureBacterial membersMembrane proteinsNutrient uptakeSequence identityMolecular insightsConformational changesHomology modelSecondary transportLeuTNeurotransmitter clearanceLipid bilayersSolute moleculesBiochemical dataFunction studiesIon bindingCrucial roleStructural models of inhibition in neurotransmitter sodium symporters
Singh S, Gouaux J. Structural models of inhibition in neurotransmitter sodium symporters. The FASEB Journal 2008, 22: 254.2-254.2. DOI: 10.1096/fasebj.22.1_supplement.254.2.Peer-Reviewed Original ResearchNeurotransmitter sodium symportersSodium symportersNovel noncompetitive inhibitorsX-ray crystallographyAtomic-level insightsSteady-state kinetic dataLeucine transporterSteady-state kineticsMolecular principlesRational designSmall moleculesIon gradientsKinetic dataDissociation assaysAddictive compoundInhibitor actionCritical roleNoncompetitive modeStructural paradigmSymporterNoncompetitive inhibitorAllosteric antagonistsGlial cytoplasmSynaptic transmissionLeuT