核酸降解与和代谢

DegradationandBiosynthesis

ofNucleotidesNucleotides,Nucleoside.Thebaseofanucleotideisjoinedcovalently:N-9ofpurinesN-1ofpyrimidines1’-carbonofthepentose,N-beta-glycosylThephosphateisesterifiedtothe5’-carbon.Twotypesofpathwaysleadtonucleotides:Thedenovoandthesalvagepathways.Denovosynthesisofnucleotidesbeginswiththemetabolicprecursors:aminoacids,ribose5-phosphate,CO2,andNH3.Salvagepathwaysrecyclethefreebasesandnucleosidesreleasedfromnucleicacidbreakdown.DNA/RNAnucleotideNucleosidephosphatebasepentoseDNases,RNasesEndonuclease,ExonucleolasephospholipaseNucleosidase:NucleosidephosphorylaseNucleosidehydrolaseDNAandRNAdegradationGenomicDNAPlamidDNADnaseIdigestionofDNARNARNAdegradationbyRnaseDNADigestionbyRestrictionEnzymes1978NOBELPRIZE证明一种特别的限制酶的存在，它只分裂那些含有为噬菌体所特有的某种序列的核苷酸。流感嗜血杆菌从噬菌体接受DNA的机制时发现了一类新的限制酶，它们分别在特定部位切断DNA分子一种限制酶只能识别一种特定的核苷酸序列，并在特定的切割点上将DNA分子切断。目前已发现的限制酶有400～500多种。DNADigestionbyRestrictionEnzymesDNAfootprint基因重组与DNA克隆（基因工程）DNADigestionbyRestrictionEnzymesAndDNAbinationPurineDegredation

DeaminationAdeninedeaminaseGuaninedeaminaseXanthineOxidaseXanthineOxidaseUrateOxidaseXanthineAdenineGuanineUricacidAllantoinAllantoinase尿酸尿囊素尿酸尿囊素尿囊酸尿素Uracil,CytosineThymineCytosineThymineUracilFinal：aminoacid,NH3,CO2PyrimidineDegradationBiogenesisofnucleotidesPentose戊糖phosphatepathwayBiogenesisofpurinesThisinformationwasobtainedfromisotopicexperimentswith14C-or15N-labeledprecursors.次黄嘌呤核苷酸Uniondeamination次黄嘌呤核苷酸腺苷琥珀酸黄嘌呤核苷酸AMPDNA/RNAnucleotideNucleosidephosphatebasepentoseDNases,RNasesEndonuclease,ExonucleolasephospholipaseNucleosidase:NucleosidephosphorylaseNucleosidehydrolaseDNAandRNAdegradationSalvagepathwayofpurinsSalvagepathwayofpurins腺嘌呤磷酸核糖转移酶APRT次黄嘌呤鸟嘌呤磷酸核糖转移酶HGPRTDefectinsalvagepathwayofpurinsLesch(/leski/)-Nyhansyndrome:NOHGPRTActivity!producedbymutationsintheHPRTgenelocatedontheXchromosomeRegulationofpurinenucleotideBiosynthesis:coordinationofdenovoandsalvagepathwayLesch(/leski/)-NyhansyndromePRPPSubstrateactivationTheGoutOverproductionofuricacidUricacidcrystalsprecipitateintojoints(GoutyArthritis痛风性关节炎),kidneys,ureters(stones结石)AdeninedeaminaseGuaninedeaminaseXanthineOxidaseAdenineSalvagepathwaydeficiency,Cancertreatment,Allopurinol:hypoxanthineanalogXanthineoxidaseinhibitorsbindstoXanthineOxidaseinhibitproductionofuricacid别嘌呤醇次黄嘌呤缺乏次黄嘌呤-鸟嘌呤磷酸核糖转移酶（HGPRT）HAT培养基含有次黄嘌呤(H)、氨基喋呤(A)和胸腺嘧啶核苷(T)具有次黄嘌呤-鸟嘌呤磷酸核糖转移酶（HGPRT）杂交瘤细胞筛选在HAT培养基[属于选择性培养基，含有次黄嘌呤(H)、氨基喋呤(A)和胸腺嘧啶核苷(T)]中进行选择性培养，未融合的脾细胞因不能在体外长期存活而死亡；未融合的骨髓瘤细胞合成DNA的主要途径被培养基中的氨基蝶呤阻断，又因缺乏次黄嘌呤-鸟嘌呤-磷酸核糖转移酶（HGPRT），不能利用培养基中的次黄嘌呤完成DNA的合成过程而死亡。只有融合的杂交瘤细胞由于从脾细胞获得了次黄嘌呤-鸟嘌呤-磷酸核糖转移酶，因此能在HAT培养基中存活和增殖。经过克隆选择，可筛选出能产生特异性单克隆抗体的杂交瘤细胞，在体内或体外培养，即可无限制地大量制备单克隆抗体。PyrimidinebiosynthesisCarbamoylphosphateAspartate门冬氨酸转氨甲酰酶二氢乳清酸酶Orotate乳清酸orotidylateDecarboxylaseCarbamoylphosphateAspartateUMPN-CarbamoylaspartateaspartatetranscarbamoylasedihydroorotaseL-Dihydroorotatedihydroorotatedehydrogenaseorotatephosphoribosyl-transferaseOrotidylateUMP合成酶缺陷：遗传性乳清酸尿症ATPADPUMPKinaseUDPUDPKinaseATPADPUTPCTPsynthetaseGlnATPGluADP+PiAMP,GMP,UMP,CMPPurinebiosynthesisPyrimidinebiosynthesisPurinesalvagepathwaysOverviewofnucleotidebiosynthesisFormationofdeoxy-NucleotidesNucleoside5-diphosphatereductaseRNAAGCUDNAAGCUADP,GDP,UDP,CDPdADP,dGDP,dUDP,dCDP

UMPtodTMP

UMP→UDPdUDP→dUMPdTMP

-(O)+CH3(THF)RNAAGCUDNAAGCUTThymidylatesynthaseleadstomethylationofdUMPandproducesdTMP.ThymidylatesynthaseisespeciallyneededforDNAreplication/repair.ItisthusatargetforcancertreatmentlikefluoruracilBiogenesisofpurinesandpyromidine叶酸InhibitstheDNAsynthesisofBacterialbutnotofhuman.DrugsinterferingbiosynthesisofDNA