不同价态金属离子对BSA结构的影响(2)

3+)/(1.0×10 6mol/L):0,1,5,10,20,40,60,100respectively;pH7.4.

10000900080007000

6000S

LR50004000300020001000

Concentration (10-6

mol/L)

Fig.2.Resonancelightscattering(RLS)peaksignalsofBSAwithdifferentconcen-trationsofionsNa+,Cu2+,Al3+.Conditions:c(BSA)=1.0×10 6mol/L,c(Na+,Cu2+,Al3+)/(1.0×10 6mol/L):0,1,10,20,40,50,60,100respectively;pH7.4.

andphenylalanine(Phe).TheresultofAl3+–BSAwassimilartoNa+–BSA.4.2.RLSspectra

Fig.2showedthecomparisonofRLSspectraaffectedby

Na+–BSA,

Cu2+–BSAandAl3+–BSA.Astrongpeakwasobservedat

272nm.WefoundthatRLSofNa+–BSAremainedstable,andthepeaksignalschangedlittle,whichindicatedtheparticleofNa+–BSAmaintainedstable.ThepeaksignalsofRLSinCu2+–BSAsystemincreasedwiththedoseofCu2+,whichillustratedthatparticlesinthissystembecamelargerbecausesomeactingforcemayworkbetweenCu2+andBSA,andthisforcewasstrengthenedwithCu2+increased.ThepeaksignalsofAl3+–BSAremainedstableinlowconcentrationofAl3+,buttheywouldincreasevisiblywhentheconcentrationofAl3+exceeded4.0×10 6mol/L.WehavetestedtheRLSofAl3+aloneinordertorecognizetherealeffectprinci-plebetweenAl3+andBSA.TheresultswereshowninTable1.1.Table1.1showedpeaksignalsofAl3+–BSAwerestrongerthanthoseofAl3+alonewhenc(Al3+)≤2.0×10 5mol/L.WhiletheresultswouldgobycontrarieswithincreasingAl3+dose,andthesubtrac-

H.Zhangetal./SpectrochimicaActaPartA78 (2011) 523–527

525

Table1.1

ThecomparisonofRLSbetweenAl3+–BSAandAl3+alone.Systems01102040

BSA+Al3+

4910

4788474050055131Al3+0242633514046168 RLS

4910

2362

1389

959

2037

Ps:CmeansconcentrationofAl3+.SignalexceedsthedetectionlimitwhenC5.0×10 5mol/L.

RLSstandsforthesubtractionbetweenthepeaksignalsofBSA+Al3+andAl3+alone.

Table1.2

ThecomparisonofRLSbetweenNa+–BSAandNa+alone.SystemsC(10 6mol/L)110100BSA+Na+

464746864854Na+416529212931 RLS

482

1765

1923

tion( RLS)wasbecomingsmaller.TheresultindicatedthatBSAwouldnotreactwithAl3+inlowconcentrationofAl3+,andthepeaksignalsofRLSremainedstable;Whenc(Al3+)≥2.0×10 5mol/L,peaksignalsofthesystemraisedwithAl3+increased,butweakerthanthoseofAl3+alone,soBSAreactedwithAl3+initshighcon-centration,andformedalargeraggregationsimultaneously.Atthesametime,hydrolysisofAl3+-binedwiththeresultof uorescenceemissionspectrainAl3+–BSAsystem,wecancometoaconclusionthatthetoxiceffectofAl3+onBSAisweakenoughtoin uencethestructureofBSAvisi-bly,onlychangingthesolutionenvironmentofBSAbyhydrolysisofAl3+.

Forthesamereason,thecomparisonsofanothertwoionswereillustratedinTables1.2and1.3aswell.Wecan gureoutthatpeaksignalsofNa+–BSAwerestrongerthanthoseofNa+aloneintheconcentrationrange,andCu2+sharesthesameresultwithNa+.SowecangetaconclusionsafelythatNa+doesnotreactwithBSA,whileCu2+couldbindwithBSAbysomeactingforce,whichaggregateslargerparticles.

Theresultwhichwecangetfrom uorescenceemissionspectraandRLSgivesusanilluminationthationelectricchargeisnotthemainfactoraffectingthestructureofBSA.4.3.Synchronous uorescencespectra

Synchronous uorescencespectroscopycangiveinformationaboutthemolecularenvironmentinthevicinityofachro-mophoresuchastryptophanandtyrosine.Theshiftintheemissionmaximum( em)re ectsthechangesofpolarityaroundthechro-mophoremolecule[19].WetestedCu2+–BSAwithsynchronous uorescencespectrainorderto ndoutthequenchingmechanisminwhichCu2+playedarolewiththe uorescenceofBSAresidues,Trp,TyrandPheinparticular.When betweenexcitationandemissionwavelengthissetat15nm,thesynchronous uorescencegivescharacteristicinformationofTyr.Andwhen is xedat60nm,aspectrumcharacteristicofTrpisobtained[20,21].TheresultsdetectedbyF-4600 uorescencespectrophotometerwereillustratedinFig.3.

Table1.3

ThecomparisonofRLSbetweenCu2+–BSAandCu2+alone.SystemsC(10 6mol/L)110100BSA+Cu2+

155817419066Cu2+77111094935 RLS

787

632

4131

y

tisnetni ecnecseroulFy

tisnetni ecnecseroulFFig.3.SynchronousFluorescenceSpectraofBSAwithdifferentconcen-trationsofcopperchloride,(A) =15nm,(B) =60nm.Conditions:c(BSA)=1.0×10 6mol/L,c(Cu2+)/(1.0×10 6mol/L):0,1,5,20,50,100respectively;pH7.4.

Wecan gureoutfromFig.3thatbothTrpandTyrwerequenchedsimultaneously,andtheintensityofTrpdecreasedsharperthanthatofTyr,whichindicatedbindingsitesbetweenCu2+andBSAwasclosertoTrp[22].WiththeconcentrationofCu2+increasedgradually,aslightredshiftofTrpandTyrpeakcouldbeobserved,whichprovedmolecularconformationofBSAchangedinthetoxiceffectofCu2+,thehydrophobicityoftheTrpandTyrdecreased,andbothoftwochromophoresburiedinthenon-polarhydrophobiccavitiesweremovedtoamorehydrophilicenvironment[23,24].

Theinteractionbetweenheavymetalsandbiomoleculesisavastresearchareathatmanyresearchershavestudiedinthis eld.LiWangetal.[25]hasinvestigatedthatNi2+hadanobvioustoxiceffectonbovinehemoglobin(BHb).Zn2+,Cd2+,Mn2+,Co2+andCr2+[26–29]havealreadybeenstudiedonthetoxicityofserumalbumin.Alloftheseresultsabovewarnusthatheavymetalmakesamajorhazardtobiomolecules.4.4.UV–visabsorptionspectra

TheultravioletabsorptionspectrainFig.4wereachievedbysubtractionofNa+,Cu2+andAl3+absorption.BSAalonehastwoabsorptionpeaksatabout200and280nm,whichstandsforthestrongabsorptionofpeptidebondandaromaticresiduesrespec-tively.AswecanseeinFig.4AandB,withtheconcentrationofNa+andAl3+increased,thepeakshapeandpositionremainedstablein

526

H.Zhangetal./SpectrochimicaActaPartA78 (2011) 523–527

2.5

A

2.0

1.5

s

bA1.0

0.5

0.0

200220240260280300320

Wavelength(nm)

2.5

B

2.0

1.5

s

bA1.0

0.5

0.0

200220240260280300320

Wavelength(nm)

s

bAWavelength(nm)

Fig.4.UV–Vis.AdsorptionspectraofBSAwithdifferentconcentrationsofsodiumchloride,copperchlorideandaluminiumchlorideConditions:c(BSA)=1.0×10 7mol/L,c(Na+,Al3+)/(1.0×10 6m …… 此处隐藏：5541字，全部文档内容请下载后查看。喜欢就下载吧 ……