生物分离工程-双语版-5教案资料.ppt

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1、生物分离工程-双语版-5Denaturation1Interfacialphenomenaandsurface-surfaceinteractions2Geometriccomplementarityandaffinityinteraction3Chromatographyretentionislargelydeterminedbyaminoacidsonpolypeptidesurface4Interestproteinmaybedenaturedandrequirerefoldingtoregainthenativestructure5Denaturationreasons(1)Comp

2、etinghydrophobicandcoulombicforcesonthecolumnaregreaterthanthosemaintainingproteinsstructure(2)Mobilephasedenaturation(3)NotasinglereasontominimizeproteindenaturationAdsorptionisotherm1Ionexchangechromatographyplayamajorroleinbiotechnologyindustry2Langmuirisotherm3Freundlichisotherm4Adsorptionchara

3、cteristicsofproteinsaffectconformationaldynamicsonsurfaceIIchromatographytechniques1Popularmethodsofbioseparation(1)ionexchange(2)gelpermeation(3)affinitychromatography(4)hydrophobicinteractionchromatography2HPLCforbioseparationinrapiddevelopment(1)rigidchromatographysupports(2)highflowandrapidsepa

4、ration(3)samecriteriaasconventionalliquidchromatographyAisoelectricpointBnetchargeatdifferentpHCmolecularweightIIchromatographytechniques3Elution(1)continuedelution(2)pHgradient(3)ionstrengthgradient4Otherapplicationofchromatographytechniques(1)antibiotics(2)vitamins(3)steroids(4)peptidesAIon-excha

5、ngechromatography1Basicnetchargetheory(1)notberetainedatpI(2)proteinsretainedonpositivelychargeanionexchangerabovepI(3)proteinsretainedonnegativelychargecationexchangebelowpI(4)afunctionalrelationshipbetweennetchargeandretentiontimeofproteins2Problems:(1)significantretentioninIECoccuratpI(2)correla

6、tionbetweennetchargeandretentionispoor(3)oppositelychargedpatchandheterogeneityaffectconformationalchangesProteinsretentionmodelRetentionfactors(1)distributionofcharge(2)numberofchargedsitesinteractingwithsupportsurfaceHPIECmodelDisplacementmodechromatography1MechanismLowaffinitycomponentsfirstlyad

7、sorbedonIEC,thenhighaffinityinterestingcomponentstodisplacetheadsorbedlowaffinitycomponents2Application(1)Ovalbumin(2)SoybeantrypsininhibitorTable5.1(3)Aminoacids(4)Peptides(5)SteroidsPurificationofmonoclon