2024
Quantitative single-molecule analysis of assembly and Ca2+-dependent disassembly of synaptotagmin oligomers on lipid bilayers
Li F, Coleman J, Redondo-Morata L, Kalyana Sundaram R, Stroeva E, Rothman J, Pincet F. Quantitative single-molecule analysis of assembly and Ca2+-dependent disassembly of synaptotagmin oligomers on lipid bilayers. Communications Biology 2024, 7: 1608. PMID: 39627539, PMCID: PMC11615320, DOI: 10.1038/s42003-024-07317-9.Peer-Reviewed Original ResearchMeSH KeywordsAnimalsCalciumLipid BilayersProtein MultimerizationSingle Molecule ImagingSynaptotagmin IConceptsSyt-1Lipid bilayerRing-like oligomersCa2+-evoked releaseSynaptotagmin-1Single-molecule imaging methodsSynaptic vesiclesBiochemical evidencePhysiological requirementsOligomerizationAnalysis of assembliesBilayerOligomersCa2+LipidAssemblyCa2Classes of oligomersMutationsVesiclesDisassemblyEvoked release
2023
Direct determination of oligomeric organization of integral membrane proteins and lipids from intact customizable bilayer
Panda A, Giska F, Duncan A, Welch A, Brown C, McAllister R, Hariharan P, Goder J, Coleman J, Ramakrishnan S, Pincet F, Guan L, Krishnakumar S, Rothman J, Gupta K. Direct determination of oligomeric organization of integral membrane proteins and lipids from intact customizable bilayer. Nature Methods 2023, 20: 891-897. PMID: 37106230, PMCID: PMC10932606, DOI: 10.1038/s41592-023-01864-5.Peer-Reviewed Original ResearchConceptsIntegral membrane proteinsMembrane proteinsOligomeric organizationOligomeric stateNative mass spectrometry analysisFunctional oligomeric stateKey membrane componentMass spectrometry analysisNMS analysisTarget membraneLipid bindingMembrane componentsProteolipid vesiclesMembrane compositionLipid compositionSpectrometry analysisLipid membranesNeurotransmitter releaseProteinMembraneLipidsMembrane propertiesDirect determinationBilayersTransporters
2019
Highly Reproducible Physiological Asymmetric Membrane with Freely Diffusing Embedded Proteins in a 3D‐Printed Microfluidic Setup
Heo P, Ramakrishnan S, Coleman J, Rothman JE, Fleury J, Pincet F. Highly Reproducible Physiological Asymmetric Membrane with Freely Diffusing Embedded Proteins in a 3D‐Printed Microfluidic Setup. Small 2019, 15: e1900725. PMID: 30977975, DOI: 10.1002/smll.201900725.Peer-Reviewed Original ResearchMeSH KeywordsDimethylpolysiloxanesFluorescence Recovery After PhotobleachingLipid BilayersMicrofluidicsPrinting, Three-DimensionalConceptsMost biological processesLipid leafletAreas of biologyEmbedded proteinsBiological processesRelevant lipidsProteinAsymmetric bilayersPhysiological conditionsModel membranesPlanar bilayersBilayer formation processInvaluable insightsBilayersConfocal microscopeMembraneLipidsTransmembraneBiologyLeafletsMicrofluidic setupRecapitulation
2018
High-Throughput Monitoring of Single Vesicle Fusion Using Freestanding Membranes and Automated Analysis
Ramakrishnan S, Gohlke A, Li F, Coleman J, Xu W, Rothman JE, Pincet F. High-Throughput Monitoring of Single Vesicle Fusion Using Freestanding Membranes and Automated Analysis. Langmuir 2018, 34: 5849-5859. PMID: 29694054, DOI: 10.1021/acs.langmuir.8b00116.Peer-Reviewed Original ResearchConceptsMembrane fusionFusion eventsSoluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteinsSNARE-dependent membrane fusionIndividual vesicle fusion eventsFactor attachment protein receptor proteinsN-ethylmaleimide-sensitive factor attachment protein receptor proteinsT-SNARE proteinsSingle-vesicle fusionProtein receptor proteinsVesicle fusion eventsMobility of proteinsVesicle dockingContent releaseVesicle fusionHigh-throughput monitoringPlanar membranesReceptor proteinLipid mixingProteinLipid bilayersVesiclesCorrect reconstitutionMembraneAqueous compartment
2016
FRAP to Characterize Molecular Diffusion and Interaction in Various Membrane Environments
Pincet F, Adrien V, Yang R, Delacotte J, Rothman JE, Urbach W, Tareste D. FRAP to Characterize Molecular Diffusion and Interaction in Various Membrane Environments. PLOS ONE 2016, 11: e0158457. PMID: 27387979, PMCID: PMC4936743, DOI: 10.1371/journal.pone.0158457.Peer-Reviewed Original ResearchConceptsFluorescence correlation spectroscopySingle-particle trackingCorresponding recovery timeFRAP measurementsDynamics of lipidsDiffusion coefficientCorrelation spectroscopyBrownian motionParticle trackingConfocal microscopeAccurate valuesDiffusive speciesMembrane environmentMolecular diffusionMeasurementsMembrane platformsBehavior of lipidsFRAP experimentsMotionA Programmable DNA Origami Platform to Organize SNAREs for Membrane Fusion
Xu W, Nathwani B, Lin C, Wang J, Karatekin E, Pincet F, Shih W, Rothman JE. A Programmable DNA Origami Platform to Organize SNAREs for Membrane Fusion. Journal Of The American Chemical Society 2016, 138: 4439-4447. PMID: 26938705, PMCID: PMC4950518, DOI: 10.1021/jacs.5b13107.Peer-Reviewed Original ResearchMeSH KeywordsDNADNA, Single-StrandedLipid BilayersLiposomesMembrane FusionProtein BindingSNARE ProteinsVesicular Transport ProteinsConceptsMembrane fusionSoluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexCore molecular machineryMembrane fusion eventsProtein receptor complexMembrane fusion processMolecular machineryDNA origami platformTarget membraneAuxiliary proteinsIntracellular communicationDocking stepSingle-event levelReceptor complexLipid mixingSmall unilamellar vesiclesLipid bilayersSnareFundamental processesVesiclesUnilamellar vesiclesTraffickingMachineryProteinFusion
2014
Calcium sensitive ring-like oligomers formed by synaptotagmin
Wang J, Bello O, Auclair SM, Wang J, Coleman J, Pincet F, Krishnakumar SS, Sindelar CV, Rothman JE. Calcium sensitive ring-like oligomers formed by synaptotagmin. Proceedings Of The National Academy Of Sciences Of The United States Of America 2014, 111: 13966-13971. PMID: 25201968, PMCID: PMC4183308, DOI: 10.1073/pnas.1415849111.Peer-Reviewed Original ResearchMeSH KeywordsCalciumHumansLipid BilayersMultiprotein ComplexesProtein Structure, TertiarySNARE ProteinsSynaptotagmin IConceptsSynaptic vesicle protein Synaptotagmin 1Cytosolic domainSoluble N-ethylmaleimide-sensitive factorN-ethylmaleimide-sensitive factorMembrane fusion machineryReceptor complex assemblyRing-like oligomersFusion machineryC2 domainComplex assemblySynaptotagmin-1Helical reconstructionFusion proceedsNovel mechanismStructural mechanismsLipid monolayersNeurotransmitter releaseAbsence of calciumPhysiological concentrationsRing formationPresence of calciumFree calcium ionsSynaptotagminCalcium influxCircular arrangement
2013
COPI buds 60-nm lipid droplets from reconstituted water–phospholipid–triacylglyceride interfaces, suggesting a tension clamp function
Thiam AR, Antonny B, Wang J, Delacotte J, Wilfling F, Walther TC, Beck R, Rothman JE, Pincet F. COPI buds 60-nm lipid droplets from reconstituted water–phospholipid–triacylglyceride interfaces, suggesting a tension clamp function. Proceedings Of The National Academy Of Sciences Of The United States Of America 2013, 110: 13244-13249. PMID: 23901109, PMCID: PMC3746913, DOI: 10.1073/pnas.1307685110.Peer-Reviewed Original ResearchMeSH KeywordsADP-Ribosylation Factor 1AnimalsCoat Protein Complex IEscherichia coliHumansHydrophobic and Hydrophilic InteractionsLipid BilayersMicroscopy, ElectronPhosphatidylcholinesPhosphatidylethanolaminesPhospholipidsProtein TransportSf9 CellsSpectrometry, FluorescenceSpodopteraSurface TensionTransport VesiclesTriglyceridesWaterConceptsCoat protein complex ILipid dropletsCoat protein complexTransport vesicle formationProtein complex ILipid droplet targetingBud vesiclesCOPI subunitsProtein complexesCoat componentsNeighboring proteinsIntracellular traffickingVesicle formationComplex IClamp functionPossible direct interactionDirect interactionPhospholipid packingUnsuitable targetsProteinBilayer membranesCoatomerMembraneOrganellesProtomers
2012
SNARE Proteins: One to Fuse and Three to Keep the Nascent Fusion Pore Open
Shi L, Shen QT, Kiel A, Wang J, Wang HW, Melia TJ, Rothman JE, Pincet F. SNARE Proteins: One to Fuse and Three to Keep the Nascent Fusion Pore Open. Science 2012, 335: 1355-1359. PMID: 22422984, PMCID: PMC3736847, DOI: 10.1126/science.1214984.Peer-Reviewed Original ResearchMeSH KeywordsAnimalsCalciumDiffusionLipid BilayersLiposomesMembrane FusionMembrane ProteinsMiceNeurotransmitter AgentsProtein Structure, TertiaryProteolipidsRatsRecombinant Fusion ProteinsSNARE ProteinsSynaptic TransmissionSynaptic VesiclesSynaptosomal-Associated Protein 25Syntaxin 1Vesicle-Associated Membrane Protein 2ConceptsVesicle-associated membrane protein 2Bilayer fusionNative transmembrane domainNascent fusion poresLipid bilayer nanodiscsMembrane protein 2Synchronous neurotransmitter releaseSNARE complexTransmembrane helicesTransmembrane domainBilayer nanodiscsFused bilayersFusion porePore opensFusion partnerBiochemical studiesProtein 2Neurotransmitter releaseNanodiscsSnareEfficient releaseSynaptic transmissionSNAREpinsFusionRelevant time scales
2011
Complexin activates and clamps SNAREpins by a common mechanism involving an intermediate energetic state
Li F, Pincet F, Perez E, Giraudo CG, Tareste D, Rothman JE. Complexin activates and clamps SNAREpins by a common mechanism involving an intermediate energetic state. Nature Structural & Molecular Biology 2011, 18: 941-946. PMID: 21785413, PMCID: PMC3736826, DOI: 10.1038/nsmb.2102.Peer-Reviewed Original Research
2009
Quantification of phase transitions of lipid mixtures from bilayer to non-bilayer structures: Model, experimental validation and implication on membrane fusion
Xu W, Pincet F. Quantification of phase transitions of lipid mixtures from bilayer to non-bilayer structures: Model, experimental validation and implication on membrane fusion. Chemistry And Physics Of Lipids 2009, 163: 280-285. PMID: 20018184, DOI: 10.1016/j.chemphyslip.2009.12.002.Peer-Reviewed Original ResearchConceptsPhase transition
2007
Confinement Free Energy of Surfaces Bearing End-Grafted Polymers in the Mushroom Regime and Local Measurement of the Polymer Density
Li F, Pincet F. Confinement Free Energy of Surfaces Bearing End-Grafted Polymers in the Mushroom Regime and Local Measurement of the Polymer Density. Langmuir 2007, 23: 12541-12548. PMID: 17988162, DOI: 10.1021/la7021374.Peer-Reviewed Original ResearchConceptsPolymer chainsMushroom regimeFree energyEnd-Grafted PolymersSurface force measurementsSingle polymer chainSurface force apparatusEnd-tethered polymer chainsInteraction free energyGrafted polymersForce apparatusPolymer densityForce profilesPolymersConfinement free energyPolymer theoryLipid bilayersIdentical surfacesMushroomlike structuresFree energy approximationForce measurementsSurfacePlanar surfaceGood precisionChainEnergetics and dynamics of SNAREpin folding across lipid bilayers
Li F, Pincet F, Perez E, Eng WS, Melia TJ, Rothman JE, Tareste D. Energetics and dynamics of SNAREpin folding across lipid bilayers. Nature Structural & Molecular Biology 2007, 14: 890-896. PMID: 17906638, DOI: 10.1038/nsmb1310.Peer-Reviewed Original ResearchMeSH KeywordsAnimalsLipid BilayersMembrane FusionMiceProtein ConformationProtein FoldingRatsSNARE ProteinsSurface PropertiesHydrophobic Forces and Hydrogen Bonds in the Adhesion between Retinoid-Coated Surfaces
Tareste D, Pincet F, Lebeau L, Perez É. Hydrophobic Forces and Hydrogen Bonds in the Adhesion between Retinoid-Coated Surfaces. Langmuir 2007, 23: 3225-3229. PMID: 17266339, DOI: 10.1021/la0629779.Peer-Reviewed Original ResearchConceptsHydrophobic forcesHydrogen bondsPure waterShort-range attractive forcesLipid layerHydrophilic headgroupLong-range attractionHydrophobic chainsTris bufferHydrophobic effectLipid monolayersApolar characterMolecular interactionsAttractive forceBondsLipid bilayersExponential forceHeadgroupsNew classForce measurementsDistance profilesWaterBufferInteractionMonolayers
2006
Transition from long- to short-lived transient pores in giant vesicles in an aqueous medium
Rodriguez N, Cribier S, Pincet F. Transition from long- to short-lived transient pores in giant vesicles in an aqueous medium. Physical Review E 2006, 74: 061902. PMID: 17280091, DOI: 10.1103/physreve.74.061902.Peer-Reviewed Original Research
2005
Spontaneous and Reversible Switch from Amphiphilic to Oil-Like Structures
Pincet F, Tareste D, Amar M, Perez É. Spontaneous and Reversible Switch from Amphiphilic to Oil-Like Structures. Physical Review Letters 2005, 95: 218101. PMID: 16384186, DOI: 10.1103/physrevlett.95.218101.Peer-Reviewed Original ResearchIndirect evidence of submicroscopic pores in giant unilamelar vesicles
Rodriguez N, Heuvingh J, Pincet F, Cribier S. Indirect evidence of submicroscopic pores in giant unilamelar vesicles. Biochimica Et Biophysica Acta 2005, 1724: 281-287. PMID: 15978732, DOI: 10.1016/j.bbagen.2005.04.028.Peer-Reviewed Original ResearchConceptsPore-like structuresGiant unilamellar vesiclesOuter leafletCell membraneTransient poresFluorescent lipidLipid distributionCell compartmentVesiclesGiant vesiclesModel systemUnilamellar vesiclesLipid redistributionGiant unilamelar vesiclesMembrane defectsLipidsMembraneFormation of poresMembrane ruptureSubmicroscopic poresLeafletsExchange of matterHemifusionFluorescent labelsOuter mediumGiant vesicles formed by gentle hydration and electroformation: A comparison by fluorescence microscopy
Rodriguez N, Pincet F, Cribier S. Giant vesicles formed by gentle hydration and electroformation: A comparison by fluorescence microscopy. Colloids And Surfaces B Biointerfaces 2005, 42: 125-130. PMID: 15833663, DOI: 10.1016/j.colsurfb.2005.01.010.Peer-Reviewed Original Research
2004
Hemifusion and fusion of giant vesicles induced by reduction of inter-membrane distance
Heuvingh J, Pincet F, Cribier S. Hemifusion and fusion of giant vesicles induced by reduction of inter-membrane distance. The European Physical Journal E 2004, 14: 269-276. PMID: 15338438, DOI: 10.1140/epje/i2003-10151-2.Peer-Reviewed Original ResearchConceptsInter-membrane distanceGiant unilamellar vesiclesInner leafletMembrane fusionOuter leafletMeans of micromanipulationCommon functionFluorescence microscopyProbe redistributionVesiclesInfluenza virus hemagglutininGiant vesiclesDNA basesModel systemHemifusionUnilamellar vesiclesActual fusionVirus hemagglutininMembraneContact zoneFusionHead groupsGood agreementProteinLeaflets
2002
Energy of Hydrogen Bonds Probed by the Adhesion of Functionalized Lipid Layers
Tareste D, Pincet F, Perez E, Rickling S, Mioskowski C, Lebeau L. Energy of Hydrogen Bonds Probed by the Adhesion of Functionalized Lipid Layers. Biophysical Journal 2002, 83: 3675-3681. PMID: 12496134, PMCID: PMC1302442, DOI: 10.1016/s0006-3495(02)75367-8.Peer-Reviewed Original ResearchConceptsHydrogen bondsIntramolecular hydrogen bondsHydrogen-bonding groupsSurface force apparatusSingle hydrogen bondHydrogen-bonding partsMonolayer of lipidsDifferent energetic contributionsForce apparatusHydrophobic interactionsInteraction energyBondsKcal/Lipid layerEnergetic contributionsTheoretical studyAdhesion energyProtein systemsBond partProtein foldingProtein stabilizationHeadgroupsWaterSurfaceComplex structure