生物化学-生化课件chapter

Chapter7

PortraitofanAllostericProteinThetransitionfromanaerobic厌氧toaerobic需氧lifewasamajorstepinevolutionbecauseituncoveredarichreservoirofenergy.Eighteentimesasmuchenergyisextractedfromglucoseinthepresenceofoxygenasinitsabsence.由厌氧呼吸到需氧呼吸,生命体从葡萄糖汲取能量增加到18倍(15倍).Vertebrateshaveevolvedtwoprincipalmechanismsforsupplyingtheircellswithacontinuousandadequateflowofoxygen.1.acirculatorysystemthatactivelydeliversoxygentocells.循环系统2.theuseofoxygen-carryingmoleculestoethelimitationimposedbythelowsolubilityofoxygeninwater.载氧分子(氧在水中溶解度低)Theoxygencarriersinvertebratesaretheproteinshemoglobin(Hb)andmyoglobin(Mb).Hb,whichiscontainedinredbloodcells,servesastheoxygencarrierinbloodandalsoplaysavitalroleinthetransportofcarbondioxideandhydrogenion.Mb,whichislocatedinmuscle,providesareservesupplyofoxygenandfacilitatethemovementofoxygenwithinmuscle.肌红蛋白可从血红蛋白接受分子氧并储藏在肌肉细胞中,当氧供应受限制时释放给细胞色素氧化酶.也可促成分子氧直接从肌细胞表面转移给线粒体.MbandHbillustratemanyimportantprinciplesofproteinconformation,dynamics,andfunction.Theirthree-dimensionalstructures,knowninatomicdetail,revealmuchabouthowproteinsfold,bindothermoleculesandintegrateinformation.折叠,结合其它分子,整合信息ThebindingofO2byHbisregulatedbyH+,CO2,andorganicphosphates(2,3-BPG).Theseregulatorsgreatlyaffecttheoxygen-bindingpropertiesofHbbybindingtositesontheproteinfarfromwhereO2isbound.H+,CO2,

2,3-BPG的结合位点与氧结合位点相距甚远,但可(通过变构作用)调控氧气与血红蛋白的结合Indeed,interactionsbetweenspatiallydistinctsites,termedallostericinteractions变构作用,occurinmanyproteins.Allostericeffectsplayacriticalroleincontrollingandintegratingmoleculareventsinbiologicalsystems.Hbisthebest-understoodallostericprotein.thediscoveryofmutantHbfirstrevealedthatdiseasecanarisefromachangeofasingleaminoacidinaprotein.TheconceptofmoleculardiseasecamefromstudiesofamutantHb.Hbhasalsobeenarichsourceofinsightintothemolecularbasisofevolution.OxygenBindstoaHeme血红素ProstheticGroupThecapacityofMborHbtobindoxygendependsonthepresenceofanonpolypeptideunit,namely,ahemegroup.ItgivesMbandHbtheirdistinctivecolor(红色含铁载氧色素).Manyproteinsrequiretightlybound,specificnonpolypeptideunitsfortheirbiologicalactivities.Suchaunitiscalledaprostheticgroup辅基.Aproteinwithoutitscharacteristicprostheticgroupistermedanapoprotein脱辅蛋白.脱辅蛋白+辅基=全蛋白Thehemeconsistsofanorganicpartandanironatom.Theorganicpart,protoporphyrin原卟啉,ismadeupoffourpyrrolerings吡咯环.血红素=原卟啉(及侧链)+铁原子原卟啉=四个吡咯环+连接它们的甲叉桥Thefourpyrrolesarelinkedbymethanebridges甲叉桥toformatetrapyrrolering.Fourmethyl甲基,twovinyl乙烯,andtwopropionate丙酸sidechainsareattached.Theironatominhemebindstothefournitrogensinthecenteroftheprotoporphyrinring.Theironcanformtwoadditionalbonds,oneoneithersideofthehemeplane.Theseironatomscanbeintheferrous(+2)ortheferric(+3)oxidationstate,andthecorrespondingformsofHbarecalledferrohemoglobin亚铁血红蛋白andferrihemoglobin高铁血红蛋白(alsocalledmethemoglobin).Onlyferrohemoglobin,the+2oxidationstate,canbindoxygen.ThesamenomenclatureappliestoMb.只有两价态的亚铁血红蛋白才能结合氧

MyoglobinHasaCompactStructureandaHighContentofαhelicesTheelucidationofthethree-dimensionalstructureofMbandHbarelandmarksinbiochemistry.x-raycrystallography结晶学canrevealthestructureofmoleculesaslargeasproteins.KendrewchoseMbforx-rayanalysisbecauseitisrelativelysmall,easilypreparedinquantity,andreadilycrystallized.肌红蛋白分子量小,易大量制备,易结晶IthadtheadditionaladvantageofbeingcloselyrelatedtoHb.与血红蛋白十分相似Mbfromtheskeletalmuscleofthespermwhale抹香鲸wasselectedbecauseitisstableandformsexcellentcrystals.Theskeletalmuscleofdivingmammals,suchaswhales,seals,andporpoises海豚,isparticularlyrichinMb,whichservesasastoreofoxygenduringadive.水生哺乳动物用肌红蛋白作为潜水时氧气储存In1957,Kendrewandhiscolleaguessawwhatnoonehaseverseenbefore:athree-dimensionalpictureofaproteinmoleculeinallitscomplexity.Thestructurewasmoreintricatethanhadbeenimagined.Itsmeaningwasrevealedtwoyearslater,whenahigh-resolutionimagewasobtained.Awealthofstructuraldetailemerged.Thepositionsof1200ofthe1260nonhydrogenatomswereclearlydefinedtoaprecisionofbetterthan0.3Ǻ.Thecourseofthemainchainandthepositionofthehemegroupareshown.SomeimportantfeaturesofMbareMbisextremelycompact.极为紧密About75%ofthemainchainisinanα-helicalconformation.8majorhelicalsegments,5nonhelicalsegmentsliebetweenhelices,twoothernonhelicalregions.富含α-螺旋(共8个,即ABCDEFGH),5个非螺旋片段居螺旋之间(BC,DE间无连接片段),另2个非螺旋片段分别在肌红蛋白的氨基端和羧基端Fourofthehelicesareterminatedbyaprolineresidue,whosefive-memberedringdoesnotfitwithinastraightstretchofαhelix.Themain-chainpeptidegroupsareplanar,andthecarbonylgroupofeachistranstotheNH.主链肽基在一个平面上,其中C=O和NH为反式关系.Theinsideandoutsidearewelldefined.Thereislittleemptyspaceinside.

内部几乎无空隙

AHindered阻隔的HemeEnvironmentisEssentialforReversibleOxygenation氧合

ThehemegroupislocatedinacreviceintheMbmolecule.Thehighlypolarpropionatesidechainsofthehemeareonthesurface.AtphysiologicalPH,thesecarboxylicacidgroupsareionized.(血红素的)离子化的丙酸侧链处在蛋白表面Therestofthehemeisinsidethemolecule,whereitissurroundedbynonpolarresiduesexceptfortwohistidines.血红素其它部分处于非极性残基包围之中(除两个组氨酸外)TheironatomofthehemeisdirectlybondedtooneoftheseHis.ThisHis,whichoccupiesthefifthcoordinationpositioniscalledtheproximalhistidine邻接(近側)组氨酸(F8).Theironatomisabout0.3Ǻoutoftheplaneoftheporphyrin卟啉,onthesamesideasHisF8.Theoxygen-bindingsiteisontheothersideofthehemeplane,atthesixthcoordinationposition配位位置.AsecondHisresidue(E7),termedthedistal远側histidine,isnearthehemebutnotbondedtoit.TheconformationsofthethreephysiologicallypertinentformsofMb—deoxymyoglobin,oxymyoglobin,andferrimyoglobin—areverysimilarexceptatthesixthcoordinationposition..TheaxisoftheboundO2isatanangletotheiron-oxygenbond.Theoxygen-bindingsitecomprisesonlyasmallfractionofthevolumeoftheMbmolecule.WhyisthepolypeptideportionofMbneededforoxygentransportandstorage?氧结合部位只占蛋白分子的极小部分,肌红蛋白的其它部分起阻隔铁—氧—铁“三明治”形成(“三明治”内的铁原子被氧化为3价)的作用.Theanswerliesintheoxygen-bindingpropertiesofanisolatedhemegroup.Inwater,afreeferroushemegroupcanbindoxygen,butitdoessoforonlyafleetingmoment稍纵即逝.ThereasonisthatO2veryrapidlyoxidizedtheferroushemetoferricheme,whichcannotbindoxygen.(heme-O2–hemesandwich)Inmyoglobin,thehemegroupismuchlesssusceptibletooxidationbecausetwomyoglobinmoleculescannotreadilyassociatetoformaheme-O2-hemecomplex.TheformationofthissandwichisstericallyhinderedbythedistalHisandotherresiduessurroundingthesixthcoordinationsite.Thestrongestevidencefortheimportanceofstericfactorsindeterminingtherateofoxidationofhemecomesfromstudiesofsyntheticmodelcompounds(picket-fence柵栏ironporphyrincomplexes).ThesecompoundshaveaprotectiveenclosureforbindingO2ononesideoftheporphyrinring,whereastheothersideisleftunhinderedsothatitcanbindabase.Infact,whenthebaseissubstitutedimidazole咪唑(likeHis),theoxygenaffinityofthepicket-fencecompoundislikethatofMb.Furthermore,thepicketfencestabilizestheferrousformofthisironporphyrinandthusenablesittoreversiblybindoxygenforlongperiods.人工合成柵栏铁卟啉化合物具有与Mb相似的氧亲和力,(柵栏)使得此铁卟啉稳定在二价态,并能长时间地克逆结合(释放)氧.与游离血红素不同.Thecriticaldifferencebetweenthismodelcompoundandfreehemeisthepresenceofthepicketfence,whichblockstheformationofthesandwichdimmer二聚体.(铁-氧—铁-氧)CarbonMonoxideBindingisDiminishedbythePresenceoftheDistalHistidineCarbonmonoxideisapoisonbecauseitcombineswithferromyoglobinandferrohemoglobinandtherebyblocksoxygentransport.AnisolatedhemeinsolutiontypicallybindsCOsome25,000timesasstronglyasO2.However,thebindingaffinityofMbandHbforCOisonlyabout200timesasgreatasforO2.血(肌)红蛋白中的血红素结合一氧化碳的亲和力比它的游离状态低得多Howdotheseproteinssuppresstheinnatepreferenceofhemeforcarbonmonoxide?Theanswercomesfromx-raycrystallographicandinfrared红外spectroscopicstudiesofcomplexesofCOandO2withisolatedironporphyrins.theproteinforcesCOtobindatanangleratherthaninline.ThisbentgeometryintheglobinsweakenstheinteractionofCOwiththeheme.ThedecreasedaffinityofMbandHbforCOisbiologicallyimportant.Carbonmonoxidewasapotentialhazardlongbeforetheemergenceofindustrializedsocieties.Thelevelofendogenouslyformed内源carbonmonoxideissuchthatabout1%ofthesitesinMbandHbareblockedbyCO,atolerabledegreeofinhibition.However,endogenouslyproducedCOwouldcausemassivepoisoningiftheaffinityoftheseproteinsforCOwerelikethatofisolatedironporphyrins.ThischallengewassolvedbytheevolutionofhemeproteinsthatdiscriminatebetweenO2andCObystericallyimposingabentandhenceweakermodeofbindingforCO.Thus,Mbhascreatedaspecialmicroenvironmentthatconfersdistinctivepropertiesonitsprostheticgroup.Ingeneral,thefunctionofaprostheticgroupismodulatedbyitspolypeptideenvironment.多肽微环境赋予其辅基独特的性质和功能Forexample,thesamehemegrouphasquiteadifferentfunctionincytochromec,aproteinintheterminaloxidationchaininthemitochondriaofallaerobicorganisms.Incytochromec细胞色素c,thehemeisareversiblecarrierofelectronsratherthanofoxygen.Hemehasyetanotherfunctionintheenzymecatalase过氧化氢酶,whereitcatalyzestheconversionofhydrogenperoxideintowaterandoxygen.

不同蛋白含有相同的辅基(血红素)→不同的多肽微环境(蛋白构象)使相同的辅基具有不同的功能→(血红蛋白输氧,细胞色素c电子载体,过氧化氢酶催化过氧化氢转变成水和氧)TheCentralExonofMyoglobinEncodesaFunctionalHeme-BindingUnit

mosteukaryoticgenesaremosaicsofexons(codingsequences)andintrons(noncodinginterveningsequences).exonsencodediscretestructuralandfunctionalunitsofproteins.分离的外显子编码各自(所对应)的结构域或功能域Myoglobinillustratesthisgeneralprinciplenicely.ThegeneforMbconsistsofthreeexons:nearlytheentireheme-bindingsiteisspecifiedbythecentralexon.Theregionsencodedbytheothertwoexonsmakeveryfewcontactswiththeheme.Digestionofapomyoglobinwithclostripain梭菌蛋白酶,anarginine-specificprotease,yieldsapolypeptidecontainingresidues32to139,whichcorrespondstothecentralexonandpartoftheC-terminalone.Thisfragmentbindsheme.Furthermore,thiscomplex,calledmini-myoglobin,bindsO2andCOreversibly.Indeed,theratesofassociationanddissociationarenearlythesameasthoseoftheintactprotein.Thesefindingsindicatethattheconformationofmini-myoglobinisverysimilartothatofnativemyoglobin.Becausetheamino-terminalexonandthelast14residuesoftheC-terminalexonarenotessentialforreversibleoxygenationitisevidentthatthecentralexoncontainsmuchoftheinformationforbindinghemeandmaintainingthenativefoldofanoxygen-bindingprotein.中央外显子含有结合血红素和保持(此氧结合)蛋白天然构象的信息

Theexon-intronorganizationoftheconstituentchainsofHbisverysimilartothatofMb.AminoacidsequencecomparisonssuggestthatthegenesforMbandHbdivergedsome700millionyearsago.Thus,thecentralexonoftheseoxygencarriersisanancientpieceofDNAthatencodedafunctionalheme-bindingmoduleeonsago.HemoglobinConsistsofFourPolypeptideChainsVertebrateHb,theoxygentransporterinerythrocytes,consistoffourpolypeptidechains,twoofonekindandtwoofanother.Thefourchainsareheldtogetherbynoncovalentattractions.Eachcontainsahemegroupandasingleoxygen-bindingsite.HbA,theprincipalHbinadults,consistsoftwoalpha(α)chainsandtwobeta(β)chains.(α2β2)AdultsalsohaveaminorHb(~2%ofthetotalHb)calledHbA2,whichcontainsdelta(δ)chainsinplaceoftheβchainsofHbA(α2δ2).EmbryosandfetuseshavedistinctiveHb.Shortlyafterconception妊娠,embryossynthesizezeta(ζ)chains(whichareα-likechains)andepsilon(ε)chains(whichareβ-like).Inthecourseofdevelopment,ζisreplacedbyα,andεisreplacedbygamma(γ),andthenbyβ.ζ→α

ε→γ→βThemajorHbduringthelattertwo-thirdsoffetallife,HbF,hasthesubunitcompositionα2γ2.Theαandζchainscontain141residues;theβ,γ,andδchainscontain146residues.WhydoesHbconsistofmultiplepolypeptides,andwhydotheydiffer?SubunitinteractionsareattheheartofHb’scapacitytotransportO2,CO2,andH+inaphysiologicallyresponsiveway.

正常成人HbA(α2β2),少数成人HbA2(α2δ2),胎儿(三个月前)

ζ2ε2,胎儿(三至九个月)HbF(α2γ2),出生后α2β2.X-RayAnalysisofHemoglobin:aLaborofLove心甘情愿的工作(乐此不疲)overaQuarterCenturyTheHbmoleculeisnearlyspherical,withadiameterof55Ǻ.Thefourchainsarepackedtogetherinatetrahedralarray.Thehemegroupsarelocatedincrevicesneartheexteriorofthemolecule,oneineachsubunit.Thefouroxygen-bindingsitesarefarapart;thedistancebetweenthetwoclosestironatomsis25Ǻ.Eachαchainisincontactwithbothβchains.Incontrast,therearefewinteractionsbetweenthetwoαchainsorbetweenthetwoβchains.

HemoglobinSubunitsCloselyResembleMyoglobininThree-DimensionalStructureThethree-dimensionalstructuresofMbandtheαandβchainsofhumanHbarestrikinglysimilar.

TheeighthelicesineachchainofHbarevirtuallysuperposable可叠合onthoseofMb.Thiscloseresemblanceinthefoldingoftheirmainchainswasunexpectedbecausetheiraminoacidsequencesareratherdifferent.Infact,thesethreechainsareidenticalatonly24of141positions.Hence,quitedifferentaminoacidsequencescanspecifyverysimilarthree-dimensionalstructures.Itisevidentthatthethree-dimensionalformofspermwhaleMbandoftheαandβchainsofhumanHbhasbroadbiologicalsignificance.Infact,thismotif模体,calledtheglobinfold珠蛋白折叠,iscommontoallknownvertebrateMb.theintricatefoldingofthepolypeptidechain,firstdiscoveredinMb,isnature’sfundamentaldesignforanoxygencarrier:itplacesthehemeinanenvironmentthatenablesittocarryoxygenreversibly.ThegenesforMbandforα,β,andotherchainsofHbarevariationsonafundamentaltheme.Thisfamilyofgenesalmostcertainlyarosebygeneduplicationanddiversification多样化.TheaminoacidsequencesofHbfrommorethan60species(rangingfromlampreyeels七鳃鳗tohumans)areknown.Acomparisonofthesesequencesshowsconsiderablevariabilityatmostpositions.However,ninepositionshavethesameresidueinmostspeciesstudiedthusfar.固定不变的(九个)残基都有特定的重要功能ThesehighlyconservedresiduesareespeciallyimportantforthefunctionoftheHbmolecule.Forexample,theinvariantF8histidineisdirectlybondedtothehemeiron.Severalofthemdirectlyaffecttheoxygen-bindingsite.Anotherinvariantresidue,tyrosineHC2(H螺旋羧基端的非螺旋区),stabilizesthestructurebyformingahydrogenbindbetweentheHandFhelices.GlycineB6isinvariantbecauseofitssmallsize:asidechainlargerthanahydrogenatomwouldnotallowtheBandEhelicestoapproacheachotherascloselyastheydo.ProlineC2maybeessentialbecauseitdefinesoneendoftheChelix.TheaminoacidresiduesintheinteriorofHbvaryconsiderably.However,thechangeisalwaysofonenonpolarresidueforanother(asfromalaninetoisoleucine).Hb球状分子内部残基(组成)变异很大,但都是非极性残基之间的替代Thus,thestrikingnonpolarcharacteroftheinteriorofthemoleculeisconserved.Thenonpolarcoreisimportantinbindingthehemegroupandinstabilizingthethree-dimensionalstructureofeachsubunit.Incontrast,surfaceresiduesarehighlyvariable.Indeed,fewareconsistentlypositivelyornegativelycharged.Itmightbethoughtthatprolineresidueswouldbepreservedbecauseoftheirroleashelixbreakers.However,thisisnotso.Onlyoneprolineisinvariant,yetthelengthsanddirectionsofthehelicesinallglobinsareverysimilar.Obviously,thereareotherwaysofterminatingorbendingαhelices.Forexample,anαhelixcanbedestabilizedbyhydrogenbindingbetweentheOHgroupofaserineorthreonineresidueandamain-chaincarbonylgroup.AllosterricInteractionsEnableHemoglobintoCoordinatelyTransportO2,CO2,andH+TheαandβsubunitsofHbhavethesamestructuraldesignasMb.However,newpropertiesofprofoundbiologicalimportanceemergewhendifferentsubunitscametogethertoformatetramer.不同亚单位组成的四聚体结构使Hb具有一些Mb所没有的,具重要生物学意义的特性HbisamuchmoreintricateandsentientmoleculethanisMb.前者复杂敏感得多HbtransportsH+andCO2inadditiontoO2.Hb能运送H+andCO2

Furthermore,theoxygen-bindingpropertiesofHbareregulatedbyinteractionsbetweenseparate,nonadjacentsites.Hb的相距甚远的(四个氧结合)位点间的相互作用能调控它的氧结合性质Hbisanallostericprotein,whereasMbisnot.Thisdifferenceisexpressedinthreeways:ThebindingofO2toHbenhancesthebindingofadditionalO2tothesameHbmolecule.Inotherwords,O2bindscooperativelytoHb.Incontrast,thebindingofO2toMbisnotcooperative.Hb与氧气的结合是协同的.Mb不是.TheaffinityofHbforoxygendependsonpH,whereasthatofMbisindependentofpH.TheCO2moleculealsoaffectstheoxygen-bindingcharacteristicsofHb.BothH+andCO2promotethereleaseofboundO2.Reciprocally,O2promotesthereleaseofboundH+andCO2.pH和CO2影响Hb与氧气的结合.theoxygenaffinityofHbisfurtherregulatedbyorganicphosphatessuchas2,3-bisphosphoglycerate(BPG).TheresultisthatHbhasaloweraffinityforoxygenthanMb.

2,3-二磷酸甘油酸也影响Hb与氧气的结合.Hb(比Mb)与氧气的亲和力较低(特别在氧分压较低时).OxygenBindsCooperativelytoHbThesaturation饱和度Yisdefinedasthefractionaloccupancyofalltheoxygen-bindingsitesinasolution.ThevalueofYcanrangefrom0(allsitesempty)to1(allsitesfilled).AplotofYversuspO2,thepartialpressureofoxygen,iscalledanoxygendissociationcurve.TheoxygendissociationcurvesofMbandHbdifferintwoways:First,foranygivenpO2YishigherforMbthanforHb.ThismeansthatMbhasahigheraffinityforoxygenthandoesHb.在氧解离(结合)曲线中,给定横坐标时,Mb的Y值大,即Mb具较高的氧亲和力

OxygenaffinitycanbecharacterizedbyaquantityP50,whichisthepartialpressureofoxygenatwhich50%ofsitesarefilled.ForMb,P50

1torr;forHb,P50

26torrs.TheseconddifferenceisthattheoxygendissociationcurveofMbishyperbolic双曲线,whereasthatofHbissigmoidal

S型.ForMbY=pO2/(pO2+P50)(hyperbola)(P158)TheHillCoefficientisaMeasureofCooperativity协同性thecurveobtainedfromtheoxygen-bindingdataforHbagreeswiththeequationderivedforthehypotheticalequilibrium希尔系数:希尔图中(当Y=0.5时)的斜率Hb(O2)n＝Hb+nO2

→logY/1―Y=nlogpO2―nlogP50Aplotoflog[Y/(1-Y)]versuslogpO2,calledaHillplot,approximatesastraightline.Itsslopenatthemidpointofthebinging(Y=0.5)iscalledtheHillcoefficient.Thevalueofnincreaseswiththedegreeofcooperativity;themaximumpossiblevalueofnisequaltothenumberofbindingsites.MbgivesaHillplotwithn=1.0,whichmeansthatO2moleculesbindindependentlyofeachother,asindicatedinequation1.Mbn=1.0,即不同分子结合氧相互独立Incontrast,theHillcoefficientof2.8forHbindicatesthatthebindingofoxygeninHbiscooperative.Bindingatonehemefacilitatesthebindingofoxygenattheotherhemeonthesametetramer.Hbn=2.8,一个血红素结合(释放)氧会促进同一四聚体中其它的血红素氧的结合(释放).Conversely,theunloadingofoxygenatonehemefacilitatestheunloadingofoxygenattheothers.Inotherwords,thehemegroupsofaHbmoleculecommunicatewitheachother.ThemechanismofcooperativebindingofoxygenbyHb,sometimescalledHbinteraction.同一血红蛋白分子中的血红素之间是相互联系的TheCooperativeBindingofOxygenMakesHbaMoreEfficientOxygenTransporterWhatisthebiologicalsignificanceofthecooperativebindingofoxygenbyHb?TheoxygensaturationofHbchangesmorerapidlywithchangeinthepartialpressureofO2thanitwouldiftheoxygen-bindingsiteswereindependentofeachother.Hb的氧饱和度随氧分压变化的速率比假定的(氧结合位点相互独立)情况快得多Assumethatthealveolar肺泡pO2is100torrs,andthatthepO2inthecapillaryofanactivemuscleis20torrs.LetP50=26torrs,andtaken=2.8.ThenYinthealveolarcapillarieswillbe0.98,andYinthemusclecapillarieswillbe0.32.TheoxygendeliveredwillbeproportionaltothedifferenceinY,whichis0.66.ΔY是血红蛋白氧运载能力的量度LetusnowmakethesamecalculationforahypotheticaloxygencarrierforwhichP50isalso26torrs,butinwhichthebindingofoxygenisnotcooperative(n=1).ThenY(alveoli)=0.79,andY(muscle)=0.43,andsothedifferenceinYisequalto0.36.n=1时ΔY=0.36,n=2.8时ΔY=0.66Thus,thecooperativebindingofoxygenbyHbenablesittodeliver1.83timesasmuchoxygenundertypicalphysiologicalconditionsasitwouldifthesiteswereindependent.H+andCO2PromotetheReleaseof

O2:theBohrEffectMbshowsnochangeinoxygenbindingoverabroadrangeofpH,nordoesCO2haveanappreciableeffect.InHb,however,acidityenhancesthereleaseofoxygen.

IncreasingtheconcentrationofCO2(atconstantpH)alsolowerstheoxygenaffinity.Inrapidlymetabolizingtissue,suchascontractingmuscle,muchCO2andacidareproduced.ThepresenceofhigherlevelsofCO2andH+inthecapillariesofsuchmetabolicallyactivetissuepromotesthereleaseofO2fromoxyhemoglobin.Thereciprocaleffectoccursinthealveolarcapillariesofthelungs.The[O2]↑thereunloadsH+andCO2fromHb.TheselinkagesbetweenthebindingofO2,H+,andCO2areknownastheBohreffect.玻尔效应:血红蛋白的氧亲和力随pH(和CO2分压)的变化而变化.BPGLowerstheOxygenAffinityofHbTheoxygenaffinityofHbwithinredcellsislowerthanthatofHbinfreesolution.2,3-BPGbindstoHbandhasalargeeffectonitsaffinityforoxygen.Thishighlyanionic阴离子organicphosphateispresentinhumanredcellsataboutthesamemolarconcentrationasHb.IntheabsenceofBPG,theP50oftheHbis1torr,likethatofMb.Initspresence,P50es26torrs.Thus,BPGlowerstheoxygenaffinityofHbbyafactorof26,whichisessentialinenablingHbtounloadoxygenintissuecapillaries.BPGdiminishestheoxygenaffinityofHbbybindingtodeoxyhemoglobinbutnottotheoxygenatedform.红细胞内存在2,3-BPG,在低(高)氧分压的条件下(通过与脱氧Hb的结合),大大降低Hb的氧亲和力(不结合,无作用).FetalHbhasaHigherOxygenAffinitythanMaternalHbFetuseshavetheirownkindofHb,calledHbF(α2γ2),whichdiffersfromadultHbA(α2β2).AnimportantpropertyofHbFisthatithasahigheroxygenaffinityunderphysiologicalconditionsthandoesHbA.胎儿HbF的氧亲和力高于成人HbA(的氧亲和力).因为它与2,3-BPG的结合较HbA弱(这一结合会大大降低蛋白的氧亲和力).ThehigheroxygenaffinityofHbFoptimizesthetransferofoxygenfromthematernaltothefetalcirculation.这有利于氧气由母体(循环系统)向胎儿(循环系统)的传送(通过胎盘循环).HbFisoxygenatedattheexpenseofHbAontheothersideoftheplacentalcirculation.Thehigheroxygenaffinityoffetalbloodwasknownformanyyears,butanunderstandingofitsbasiscouldcomeonlyafterthediscoveryofBPG.HbFbindsBPGlessstronglythandoesHbAandconsequentlyhasahigheroxygenaffinity.IntheabsenceofHbBPG,theoxygenaffinityofHbFisactuallylowerthanthatofHbA.Weseehereaclear-cutbiologicaladvantageofhavingdistinctformsofaprotein,calledisoformsorisotypes同工型,indifferenttissues.TheQuaternaryStructureofHbChangesMarkedlyonOxygenationHbcanbedissociatedintoconstituentchains.ThepropertiesoftheisolatedαchainareverymuchlikethoseofMb.Theαchainbyitselfhasahighoxygenaffinity,ahyperbolicoxygendissociationcurve,andoxygen-bindingcharacteristicsthatareinsensitivetopH,CO2concentration,andBPGlevel.α,β4,Mb之间的氧结合性质非常相似.Theisolatedβchainsreadilyassociatetoformatetramer(β4).LiketheαchainandMb,β4lackstheallostericpropertiesofHbandhasahighoxygenaffinity.Inshort,theallostericpropertiesofHbarisefrominteractionsbetweenitssubunits.ThefunctionalunitofHbisatetramerconsistingoftwokindsofpolypeptidechains.

Hb的变构性质来自其亚基之间的相互作用.它的功能单位是两种不同肽链组成的四聚体.crystalsofdeoxyhemoglobinshatteredwhentheywereexposedtooxygen.Deoxymyoglobincrystals,incontrast,canbindandreleaseoxygenwithoutlosingtheirform.脱氧Hb的晶体在遇氧时破碎,而脱氧肌红蛋白的晶体在结合和释放氧气时晶形不变,说明Hb结合氧后发生了重大的构象的改变.TheshatteringofHbcrystalssuggestedthattheproteinundergoesamajorconformationalchangeonbindingO2.oxy-anddeoxyhemoglobindiffermarkedlyinquaternarystructure.Theoxygenatedmoleculeismorecompact.Thechangesinthecontactsbetweentheαandβchainsareofspecialinterest.应特别关注α和β链之间的界面(在氧合时四级结构的)改变

Inthetransitionfromoxy-todeoxyemoglobin,largestructuralchangestakeplaceattwoofthefourcontactregions(theα1β2contactandtheidenticalα2β1contact)butnotattheothers(theα1β1contactandtheidenticalα2β2contact).Infact,theα1β2contactregionisdesignedtoactasaswitchbetweentwoalternativestructures.Thetwoformsofthisdovetailed带鸠尾槽的interfacearestabilizedbydifferentsetsofhydrogenbonds.Thisinterfaceiscloselyconnectedtothehemegroups,andsostructuralchangesinitaffectthehemes.Reciprocally,structuralchangesatthehemesaffectthisinterface.(亚基之间的)界面结构改变和血红素(与氧结合的状态)相互影响Mostresiduesinitarethesameinallspeciesbecauseofitskeyroleinmediatingallostericinteractions.构成不同物种的(亚基之间的)界面的大多数