分子生物學(xué)：Section A 細(xì)胞與大分子

Awonderfulmolecularworld!分子生物學(xué)重要單詞

(1/5)molecularbiologystructurefunctioncentraldogmaDNAreplicationtranscriptiontranslationmechanismDNArecombinationevolution分子生物學(xué)結(jié)構(gòu)功能中心法則DNA復(fù)制轉(zhuǎn)錄轉(zhuǎn)譯機(jī)理重組進(jìn)化分子生物學(xué)重要單詞(2/5)aminoacidproteinorganismcompositionconformationdynamicsaminogroupcarboxylgroupcarbonylgroupsidechain氨基酸蛋白質(zhì)生物組成構(gòu)象動(dòng)力學(xué)氨基羧基羰基側(cè)鏈分子生物學(xué)重要單詞(3/5)hydrophobichydrophilicacidicbasicnegativechargepositivechargepeptidebondpartialdoublebondN-terminusC-terminus疏水的親水的酸性的堿性的負(fù)電荷正電荷肽鍵部分雙鍵氨基末端羧基末端分子生物學(xué)重要單詞(4/5)freerotationpolymerproteinfoldingprimarystructuresecondarystructuretertiarystructurequaternarystructureα

helixβ

sheethydrogenbond自由旋轉(zhuǎn)聚合物蛋白質(zhì)折疊一級(jí)結(jié)構(gòu)二級(jí)結(jié)構(gòu)三級(jí)結(jié)構(gòu)四級(jí)結(jié)構(gòu)α螺旋β折疊氫鍵SectionA細(xì)胞與大分子原核細(xì)胞真核細(xì)胞ProteinDNARNAA1細(xì)胞分類ProkaryoticcellEukaryoticcell根據(jù)細(xì)胞類型劃分有機(jī)體Prokaryotes（原核生物）(Simplestlivingcells)Eukaryotes（真核生物）Eubacteria（真細(xì)菌）Archaea（古細(xì)菌）Structuralfeatures1-10mm,nodistinctsubcellularorganelles,pili(纖毛),flagella(鞭毛)10-100mm,Organelles(細(xì)胞器)BiochemistryrRNAmoleculesaredifferentamongthesekingdoms(rRNAbasedphylogeny)Esterlinkage（酯鍵）Etherlinkage（醚鍵）Esterlinkage（酯鍵）Energyproduction,metabolismReplication,transcriptionandtranslationDifferentiationFormationofsporesEmbryoniccelldifferentiation重點(diǎn)PhylogenetictreedeterminedbyrRNAsequencecomparisons(rRNA序列比較得到的系統(tǒng)發(fā)育樹）A1Cellularclassificationback典型原核細(xì)胞示意圖(類核)(纖毛)(鞭毛)A1Cellularclassification(核糖體)Cellwall:topreventcelllysis細(xì)胞[菌]溶解

inenvironmentsoflowosmolarity低滲溶液Plasmamembrane:lipidbilayerandembedded植入的proteinsforsmallmoleculeexchangeGeneticmaterials:nucleiod(類核體singleandcircularchromosome),plasmid質(zhì)粒Ribosmes:proteinsynthesismachineryPili纖毛:toallowthecelltoattachtoothercellsandsurfaceFlagella鞭毛:cellmovement典型真核細(xì)胞示意圖A1CellularclassificationbackCytoskeletalFiber細(xì)胞骨架纖維：ControlstheshapeandmovementofthecellOrganizessomemetabolic

functionsDefinition:Thedaughtercellschangetheirpatternsofgeneexpressiontobecomefunctionallydifferentfromtheparentcellaftercelldivision.Themainmolecularreason:changeofthegenesbeingtranscribed,butnotthatoftheDNAcontent.Regulatedby

developmentalcontrolgenes,mutationsinthesegenesresultinabnormalbodyplans.A1CellularclassificationbackCellulardifferentiation(細(xì)胞分化)ForexampleSporeformation

芽孢形成amongprokaryotesandlowereukaryotesEmbryoniccelldifferentiate胚細(xì)胞分化intohighlyspecializedcellsamonghighereukaryotes.CrosswithCellBiologyCourseA2亞細(xì)胞器

Nucleus細(xì)胞核mitochondriaandchloroplasts,endoplasmicreticulum內(nèi)質(zhì)網(wǎng),microbodies微體,organelleisolation細(xì)胞器的分離CrosswithCellBiology（核）DNAreplication

RNAtranscription&processing

Ribosomeassembly A2Subcellularorganelles（線粒體）Mainfunction:cellularrespiration/ATPproductionviaoxidativephosphorylationDNAreplication,RNAtranscription,proteinsynthesisNuclear-encodedMitochonrialproteins:synthesisandtransportationA2Subcellularorganelles（類囊體）（基粒）Photosynthesis,

thelight-dependentassimilationofCO2andH2OtoformcarbohydratesandO2

DNAreplication,RNAtranscription,proteinsynthesisAplantspecificorganelleA2Subcellularorganelles（葉綠體）Nuclear-encodedchloroplastproteins:synthesisandtransportationdrugoxidationanddetoxificatin(內(nèi)質(zhì)網(wǎng))Proteinsynthesis蛋白質(zhì)合成（酯類代謝）（藥物的氧化和解毒）A2SubcellularorganellesMicrobodies(lysosomes,peroxisomesandglycoxysomes)（溶酶體，過氧化物酶體，乙醛酸循環(huán)體）Glyoxysomes（乙醛酸循環(huán)體）

arespecializedplantperoxisomeswhichcarryoutthereactionsofglyoxylatecycle（乙醛酸循環(huán)）.A2SubcellularorganellesOrganelleisolation:centrifugation(離心)DensitygradientcentrifugationRatezonal(velocity)centrifugation(速度區(qū)帶離心)Equilibriumcentrifugation(等密度梯度離心)Differential(speed)centrifugation:A2Subcellularorganelles兩者差異？例、蛋白質(zhì)和核酸的分離和純化A3生物大分子

Protein(SectionB)Nucleicacids(SectionC)Polysaccharides

LipidsComplexmacromoleculesMBcourse(X),

Biochemistry()

Immunology()A3MacromoleculesPolysaccharidesarepolymersofsimplesugarscovalentlylinkedbyglycosidicbonds(糖苷鍵).碘和淀粉的顯色除吸附原因外，主要是由于生成包合物的緣故。什么是包合物呢?直鏈淀粉是由α-葡萄糖分子縮合而成螺旋狀的長(zhǎng)長(zhǎng)的螺旋體，每個(gè)葡萄糖單元都仍有羥基暴露在螺旋外。碘分子跟這些羥基作用，使碘分子嵌入淀粉螺旋體的軸心部位，生成包合物。關(guān)于淀粉遇碘變藍(lán)？包合物中，每個(gè)碘分子跟6個(gè)葡萄糖單元配合，淀粉鏈以直徑1.3nm繞成螺旋狀，碘分子處在螺旋的軸心部位。淀粉跟碘生成的包合物的顏色，跟淀粉的聚合度或相對(duì)分子質(zhì)量有關(guān)。隨分子質(zhì)量的增加，包合物的顏色的變化由無色、橙色、淡紅、紫色到藍(lán)色。A3MacromoleculesLipids:individuallipidsarenotstrictlymacromolecules,largelipidmoleculesarebuiltupfromsmallmonomericunitsandinvolvedinmanymacromoleculeassemblyTriglycerides(甘油三酯)Complexmacromolecules(復(fù)雜大分子）Covalent(共價(jià))ornoncovalentassociationsofmorethanonemajorclassesoflargebiomoleculeswhichgreatlyincreasesthefunctionalityorstructuralcapabilitiesofthecomplex.Nulceoprotein:nucleicacids+protein核蛋白

A3MacromoleculesGlycoprotein:糖蛋白carbohydrate+proteinLipoprotein:脂蛋白Lipid+proteinA4大分子的組裝

Proteincomplexes(蛋白質(zhì)復(fù)合體)Nucleoprotein(核蛋白)Membranes(膜)Noncovalentinteractions非共價(jià)相互作用proteincomplexes蛋白質(zhì)復(fù)合體A4LargemacromolecularAssembliesNoncovalentinteraction被運(yùn)送的貨物綁在中央的頭部區(qū)域，兩條腿附著在微管上。在驅(qū)動(dòng)蛋白的催化功能域發(fā)生的反應(yīng)提供了雙腿移動(dòng)的能量。Crystalstructureof50SrRNAA4LargemacromolecularAssembliesNulceoprotein核蛋白(1)

associationsofnucleicacidsandproteinRibosome核糖體:ribosomalproteins＋rRNAsChromatin(染色質(zhì)):Viruses:Telomerase

(端粒酶)

:RibonucleaseP：A4LargemacromolecularAssembliesdeoxyribonucleoproteinconsistingofDNA&histonestoformarepeatingunitcallednucleosomeproteincapsid衣殼+RNAorDNAreplicatingtheendsofeukaryoticchromosomes.RNAactsasthereplicationtemplate,andproteincatalyzes(催化)thereactiontRNAmaturation.Protein＋PRNANulceoprotein核蛋白(2)Plasmamembrane質(zhì)膜Noncovalent非共價(jià)

interactionsTheforceofmacromoleculeassembly大分子組裝的力A4LargemacromolecularAssembliesvanderWaalsforces:

noncovalentassociationsbetweenunchargedmolecules.

*Hydrogenbonds(dipole-dipole):nucleicacids

*Hydrophobic疏水的interaction:proteins Charge-chargeinteractions(saltbridges):chargedmoleculesCharge-dipole偶極,dipole-dipole:eitherorbothofthe participantsisadipole.Dispersion色散力

interactions:non-poplarmolecule瞬時(shí)偶極(偶極)(unchargedmolecules)B蛋白質(zhì)結(jié)構(gòu)1.1ProteinComposition1.2ProteinConformations1.3ProteinStructureandFunction:AFewExamples1.4TheDynamicsofProteins1.5Experiments1.1蛋白質(zhì)的組成1.2蛋白質(zhì)的構(gòu)象1.3蛋白質(zhì)的結(jié)構(gòu)與功能：幾個(gè)例子1.4蛋白質(zhì)動(dòng)力學(xué)1.5實(shí)驗(yàn)研究Proteinsarevitaltolife對(duì)生命而言蛋白質(zhì)至關(guān)重要Lifeismostdirectlytheworkofproteins.Proteinsalloworganismstogrowandreproduce.Theyprovideshape,strengthandmovement.Inthecell,proteinsareeverywhereanddoalmosteverything.生命幾乎就是蛋白質(zhì)的杰作。蛋白質(zhì)讓生物可以生長(zhǎng)和繁殖。它們?yōu)樯锾峁┝送庑魏土α?，以及運(yùn)動(dòng)功能。在細(xì)胞中，蛋白質(zhì)無處不在，行使著幾乎任何功能。B2 Proteinstructure－SizesAfewthousandsDaltons(×103)tomorethan5millionDaltons(×106)Someproteinscontainboundnonproteinmaterials(prostheticgroups輔基orothermacromolecules),whichaccountsfortheincreasedsizesandfunctionalitiesoftheproteincomplexs.Globularproteins:enzymesComplementaryfitofasubstratemoleculetothecatalyticsite(groove-like)onanenzymemolecule.

B2 Proteinstructure－Shapeschymotrypsin(糜蛋白酶)Fibrousproteins:importantstructuralproteins(silkfibroin,keratininhairandwools)Keratin(角蛋白)Protofibril

(初原纖維)microfibril

(微管)1.1ProteinComposition/蛋白質(zhì)的組成Sidechain—COOHH2N—H氨基羧基側(cè)鏈三個(gè)基團(tuán)總是相同，不同的氨基酸具有不同的側(cè)鏈R基團(tuán)。氨基酸A1

→Proteinsarecomposedof20kindsofaminoacids

蛋白質(zhì)是由20種氨基酸組成的96231.“Acidic”aminoacids:containingadditionalcarboxyl羧基

groupswhichareusuallyionizedCanformsaltbridges,arehydrophilic(親水)asparticacid(Asp,D，天冬氨酸)glutamicacid(Glu,E，谷氨酸)B1Aminoacids-charged2.“Basic”aminoacids:containingpositivelychargedgroupsLysine(Lys,K,賴氨酸)Theimidazolegroup(咪唑基)hasapKanearneutrality.Arginine(arg,R,精氨酸)deaguanidinogroup(胍基）Histidine(His,H，組氨酸)3.polaruncharged：Containgroupsthatformhydrogenbondswithwater,

hydrophilic親水的Asparagine(Asn,N，天冬酰氨)Containhydroxyl羥基groups.Serine(Ser,S，絲氨酸)Threonine(Thr,T，蘇氨酸)Glutamine(Gln,Q，谷氨酰氨)B1Aminoacids-polarunchargedCysteine(Cys,C，半胱氨酸)hasathiol(巰醇)group，whichisoftenoxidizestocystine胱氨酸x-S-S-xGlycine(Gly,G，甘氨酸)Proline(Pro,P，脯氨酸):iminoacid(亞氨基酸)Methionine(Met,M，甲硫氨酸):containsasulfuratom4.Aminoacids-nonpolaraliphatic脂肪族的(hydrophobic疏水)Alkyl(烷基)sidechainsAlanine(Ala,A，丙氨酸)Valine(Val,V，纈氨酸)Leucine(Leu,L，亮氨酸)Isoleucine(Ile,I，異亮氨酸)Phenylalanine(Phe,F，苯丙氨酸)Tyrosine(Tyr,Y，酪氨酸)Tryptophan(Trp,W，色氨酸)5.aromatic芳香族Aminoacids：AccountsformostofUVabsorbanceofproteinsat280nm，hydrophobic（疏水的)A2

→Aminoacidsareconnectedbypeptidebonds氨基酸是由肽鍵連接在一起的Partialdoublebond/部分雙鍵Partialdoublebond由于連接碳和氧的雙鍵也在同一個(gè)碳和鄰近的氮之間進(jìn)行分配，電子密度的重新分布使得碳-氮鍵具有部分雙鍵的特性。碳-氮鍵是肽鍵的核心成分，這種部分雙鍵特性防止了肽鍵發(fā)生自由旋轉(zhuǎn)。Rigidandflexiblebonds3Dstructure位于氨基酸里面的其它鍵是可以發(fā)生旋轉(zhuǎn)的，因此氨基酸還是容易彎曲的。線性的蛋白質(zhì)分子會(huì)發(fā)生彎曲、折疊和扭曲從而形成復(fù)雜的三維結(jié)構(gòu)N-terminusandC-terminus/N-末端和C-末端在細(xì)胞中通常C-末端帶有負(fù)電荷在細(xì)胞中通常N-末端帶有正電荷1.2ProteinConformations/1.2蛋白質(zhì)的構(gòu)象1.2.1DescribingProteinStructure1.2.2ChemicalandPhysicalBasisforProteinFolding1.2.1描述蛋白質(zhì)的結(jié)構(gòu)1.2.2蛋白質(zhì)折疊的化學(xué)和物理基礎(chǔ)1.2.1DescribingProteinStructureβsheetαhelixPrimarystructureTertiarystructureSecondarystructureQuaternarystructurePrimarystructureSecondarystructure一級(jí)結(jié)構(gòu)指多肽鏈中氨基酸的順序二級(jí)結(jié)構(gòu)指多肽中的區(qū)域通過折疊產(chǎn)生的最基本的結(jié)構(gòu)形式。四級(jí)結(jié)構(gòu)是由幾條多肽鏈組成的蛋白質(zhì)形狀。三級(jí)結(jié)構(gòu)是由不同二級(jí)結(jié)構(gòu)組成的整條多肽鏈的更復(fù)雜的構(gòu)象。αhelix/α螺旋Hydrogenbondα螺旋是多肽鏈的螺旋形彎曲彎曲中的一個(gè)N-H距離它3個(gè)氨基酸的羰基上的O發(fā)生相互作用，稱為氫鍵。螺旋是靠鏈內(nèi)氫鍵維持的。ProteinsmainlywithαhelicesHemoglobin/血紅蛋白由兩條α鏈和兩條β鏈組成Keratin/角蛋白由處于α-螺旋或β-折疊構(gòu)象的平行的多肽鏈組成βsheet/β折疊Hydrogenbondβ折疊相對(duì)扁平，氨基酸之間的結(jié)合與α螺旋相同。但發(fā)生相互作用的兩個(gè)原子間的距離可以大大超過多肽鏈中的三個(gè)氨基酸，這種作用發(fā)生在多肽鏈中大致保持相互平行的不同區(qū)段之間。Parallelandantiparallelβsheets(b)Antiparallel(a)Parallel多肽鏈中一個(gè)區(qū)段自身往返折疊數(shù)次ProteinsmainlywithβsheetsFibroin絲心蛋白（β片層結(jié)構(gòu)）Tertiarystructure/三級(jí)結(jié)構(gòu)αhelixβsheetTurn多數(shù)蛋白質(zhì)由α螺旋和β折疊兩種結(jié)構(gòu)混合而成，在一起以復(fù)雜的方式折疊產(chǎn)生更大的三級(jí)結(jié)構(gòu)。Tertiarystructure

ofmyoglobin

肌紅蛋白的三級(jí)結(jié)構(gòu)Myoglobin:aproteinthatcarriesoxygeninmuscle.肌紅蛋白：肌肉中運(yùn)載氧的蛋白質(zhì)。由一條肽鏈和一個(gè)血紅素輔基組成的結(jié)合蛋白。三級(jí)結(jié)構(gòu)，有8段α螺旋區(qū)。Quaternarystructure/四級(jí)結(jié)構(gòu)Onesubunit每條多肽鏈都有各自的三級(jí)結(jié)構(gòu)Quaternarystructureofimmunoglobulin

免疫球蛋白的四級(jí)結(jié)構(gòu)LLHHOnesubunit由兩條相同的輕鏈和兩條相同的重鏈共四條肽鏈所組成ELISA酶聯(lián)免疫吸附測(cè)定QuestionsQ1

→Whatforceshelptokeepthetertiarystructures?什么力幫助維持三級(jí)結(jié)構(gòu)？Q2

→Whatfunctionsdoproteinshave?

蛋白質(zhì)有哪些功能？Q3

→Areproteinstructuresfixed?

蛋白質(zhì)結(jié)構(gòu)是固定的嗎？Q4

→Whatdeterminestheprotein’stertiarystructure?

什么決定了蛋白質(zhì)的三級(jí)結(jié)構(gòu)？Vocabulary(1/2)covalentbondhydrogenbondionicbondvanderWaalsforcehydrophobicinteractionsulfhydrylgroupdisulfidebondpartialpositivechargenon-polarmoleculecarbonicanhydrase共價(jià)鍵氫鍵離子鍵范德華力疏水相互作用巰基二硫鍵部分正電荷非極性分子碳酸酐酶Vocabulary(2/2)actinactinfilamentkinesininduced-fitmodellock-and-keymodelATPsynthetaseallostericregulationlacrepressorlactose肌動(dòng)蛋白肌動(dòng)蛋白絲驅(qū)動(dòng)蛋白誘導(dǎo)-契合模型鎖-鑰模型ATP合成酶別構(gòu)調(diào)節(jié)lac阻遏蛋白乳糖Q1

→Whatforceshelptokeepthetertiarystructures?什么力幫助維持三級(jí)結(jié)構(gòu)？HydrogenbondsHexokinase/己糖激酶Physicalinteractionsinvolvedinproteinfolding1)Covalentbonding2)Hydrogenbonding3)Ionicbonding4)VanderWaalsforces5)Hydrophobicinteraction1.2.2ChemicalandPhysicalBasisfor

ProteinFolding1)Covalentbond/共價(jià)鍵SharedelectronsH－HH+1H+1+1+1Covalentbond:

Abondbetweentwoatomsinwhichelectronsareshared.共價(jià)鍵：兩個(gè)原子共享電子形成的鍵。Covalentbondsinproteins

蛋白質(zhì)中的共價(jià)鍵Covalentbond氨基酸的初級(jí)結(jié)構(gòu)Disulfidebond/二硫鍵Covalentbond二硫鍵氫鍵發(fā)生在與有電負(fù)性的O或N原子連接的H原子上，O或N會(huì)改變H原子的電子密度，使H原子帶部分正電荷。2)Hydrogenbond/氫鍵Hydrogenbond:

Relativelyweakinteratomicbondinvolvinghydrogenatomsthatarepartiallypositivelycharged.氫鍵：相對(duì)較弱的原子之間形成的鍵，由于氫原子上帶有部分正電荷而引起。HydrogenbondPartialchargesvs.FullchargesClδ–δ+δ+Na部分電荷完全電荷Hydrogenbondsinproteins/蛋白質(zhì)中的氫鍵Hydrogenbond氫鍵雖弱，但數(shù)量巨大，對(duì)蛋白質(zhì)的折疊產(chǎn)生很大影響。3)Ionicbond/離子鍵Theionicbondofsodiumchloride,NaClNa+11Cl+17Ionicbonds:

Anattractionbetweentwoionsofoppositecharge.離子鍵：帶相反電荷的兩個(gè)離子之間的引力。Ionicbondsinproteins/蛋白質(zhì)中的離子鍵GluLys4)VanderWaalsforces/范德華力Atom1Atom2vanderWaalsforceVanderWaalsforces:

Weakattractionscausedbyshiftsintheelectroncloudsaroundatoms.范德華力：由圍繞原子的電子云發(fā)生變化產(chǎn)生的弱引力。(1837-1923)TheNetherlandsVanderWaalsforces

vs.

IonicbondAtom1Atom2VanderWaalsforcesTheatomitselfhasnegativeandpositivepartsTheinteractionis

transientIonicbondThewholeatomiseithernegativeorpositiveTheinteractionis

stableAtom1Atom2VanderWaalsforces

vs.

hydrogenbondAtom1Atom2VanderWaalsforcesTheatomitselfhasnegativeandpositivepartsTheinteractionis

transientHydrogenbondThewholeatomiseitherpartially

negativeor

positiveTheinteractionis

stableAtom1Atom2δ–δ+δ–δ+5)Hydrophobicinteraction/疏水相互作用Hydrophobicinteraction:

Interactionthatoccursbetweennon-polarmoleculesinthepresenceofwater.疏水相互作用：當(dāng)非極性分子被置于水中發(fā)生的相互作用。?NonpolarmoleculesareforcedtogetherWatermoleculesgetcontactswitheachother.Hydrophobicinteractionsinproteins

蛋白質(zhì)中的疏水相互作用非極性氨基酸被擠到了蛋白質(zhì)的中心A1

→Fivetypesofforceshelptokeepthetertiarystructures五種力幫助維持三級(jí)結(jié)構(gòu)CovalentbondvanderWaalsforceA2

→Proteinfunctionsaregreatlydiversified蛋白質(zhì)功能極其多樣Carbonicanhydrase–Catalysis（催化）Actin–Support（支撐）Kinesin–Movement（運(yùn)動(dòng)）------Whocangivemoreexamplesofdifferentproteinfunctions?Q

→Areproteinstructuresfixed?

蛋白質(zhì)結(jié)構(gòu)是固定的嗎？1.4TheDynamicsofProteinsHexokinase/己糖激酶Glucose葡萄糖與該酶的結(jié)合會(huì)引起蛋白結(jié)構(gòu)的變化，這樣使葡萄糖分子接近ATP的末端磷酸基團(tuán)。Induced-fitvs.lock-and-keymodels活性位點(diǎn)的結(jié)構(gòu)不發(fā)生變化，這種催化機(jī)理稱為鎖-鑰機(jī)理由于底物的結(jié)合引起較大結(jié)構(gòu)變化從而促進(jìn)催化作用的機(jī)理稱誘導(dǎo)-契合機(jī)理酶-底物復(fù)合體Allostericregulation/別構(gòu)調(diào)節(jié)指某些蛋白質(zhì)的活性會(huì)因?yàn)榻Y(jié)合到它的調(diào)控位點(diǎn)上的小分子而發(fā)生改變Theprimarystructuredeterminesprotein’stertiarystructure?

一級(jí)結(jié)構(gòu)決定了蛋白質(zhì)的三級(jí)結(jié)構(gòu)TheAnfinsenExperiment/安芬森實(shí)驗(yàn)B2 Proteinstructure－motifs,DomainsandfamiliesStructuralmotifs（基序）:

GroupingsofsecondarystructuralelementsthatfrequentlyoccuringlobularproteinsOftenhavefunctionalsignificanceandrepresenttheessentialpartsofbindingorcatalyticsitesconservedamongaproteinfamilybabmotifDomains（結(jié)構(gòu)域）:

structurallyindependentunitsofmanyproteins,connectedbysectionswithlimitedhigherorderstructurewithinthesamepolypeptide.TheycanalsohavespecificfunctionsuchassubstratebindingProteinfamilies（家族）:

structurallyandfunctionallyrelatedproteinsfromdifferentsourcesTheprimarystructuresofc-typecytochromesfromdifferentorganismsMotifB3 Proteinanalysis1.Purification:toobtainenoughpuresampleforstudy2.Sequencing:determinetheprimarystructureofapureproteinsample3.Massdetermination:determinethemolecularweight(MW)ofan interestedprotein.4.X-raycrystallographyandNMR:determinethetertiarystructureofagivensample.ProteinpurificationAnexperimentalsteptopurifytheinterestedproteinfromotherproteinsandnonproteinmoleculesexistinginthecellsAnessentialexperimentalstepwhenstudyanyindividualproteinTheprincipalpropertiesofproteinsusedforpurificationSize:gelfiltrationchromatography（層析）2.Charge:ion-exchangechromatography,isoelectricfocusingelectrophoresis3.Hydrophobicity:hydrophobicinteractionchromatography4.Affinity:affinity親合