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小分子靶标与其核酸适配体亲和力的表征方法.docx

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小分子靶标与其核酸适配体亲和力的表征方法 DNA/RNAaptamershavebeenwidelyusedasmolecularprobesorinhibitorsforsmall-moleculetargetsduetotheirhighbindingaffinity,selectivity,andspecificity.However,theeffectivecharacterizationoftheaptamer-targetinteractionremainsachallengeindrugdiscovery.Inthisarticle,webrieflyreviewthecurrentmethodsforcharacterizingtheaptamer-targetinteraction,andfocusontheapplicationofisothermaltitrationcalorimetry(ITC)andsurfaceplasmonresonance(SPR)technologyinquantifyingtheaffinitybetweensmall-moleculetargetsandtheirnucleicacidaptamers. Isothermaltitrationcalorimetry(ITC)isapowerfulandusefulanalyticalandthermodynamictoolformeasuringthebindingaffinityofbiomolecules.ITCdirectlymeasurestheheatreleasedorabsorbedwhenthebiomoleculebindstoitsligand,enablingthedeterminationofthebindingconstant(Kd),stoichiometry(n),enthalpy(ΔH)andentropy(ΔS)oftheinteraction.TheprincipleofITCisbasedonthethermodynamiccharacteristicsofthereaction,namelythechangeinheattransferthataccompaniesthebindingoftwomolecules.Theenthalpychangereflectstheaffinityandspecificityoftheinteraction,whiletheentropychangereflectstherandomnessofthesystem. TheprincipleofSPRisbasedonthedetectionofchangesintherefractiveindexduringbiomolecularinteractions.Surfaceplasmonresonance(SPR)isanopticaltechniquethatcandetectandquantifybiomolecularinteractionsinreal-time,withouttheneedforlabelingoralteringthesample.SPRisoneofthemostcommonlyusedmethodsforstudyingbiomolecularinteractions,includingsmall-moleculetargetsandnucleicacids.TheadvantagesofSPRincludeitsabilitytomeasurethekineticsandaffinityofthebiomolecularinteraction,aswellastheabilitytodetectbindingeventsinreal-time. ThereareseveralimportantfactorstoconsiderwhenusingITCandSPRtostudytheinteractionbetweensmall-moleculetargetsandnucleicacidaptamers.ThefirstfactortoconsiderinITCisthepurityofthesamples,sinceimpuritiesoraggregatescaninterferewiththeaccuracyofthethermodynamicresults.Additionally,thetemperatureoftheexperimentshouldbewell-controlledandcalibrated,sincetemperaturecandire