醫(yī)學(xué)分子生物學(xué)-第二章 基因

醫(yī)學(xué)分子生物學(xué)第二章基因結(jié)構(gòu)與功能易光輝南華大學(xué)醫(yī)學(xué)院分子生物學(xué)研究中心2009-11Genes:

StructureandFunctionbyProf.GuanghuiYighyi6108@163.constituteofCardiovascularDiseaseResearchCenterforMolecularBiologyMedicalCollege,UniversityofSouthChina2009-11基因的概念及其含義的演變參見(jiàn)“醫(yī)學(xué)分子生物學(xué)---第一章緒論”相關(guān)學(xué)習(xí)網(wǎng)站：(NHGRI官方網(wǎng)站)EukaryoticCentralDogma翻譯非翻譯序列蛋白成熟RNARNA初始轉(zhuǎn)錄本外顯子內(nèi)含子RNA剪接轉(zhuǎn)錄啟動(dòng)子上游下游Gene:ADNAsegmentcontainingbiologicalinformationandhencecodingforanRNAand/orpolypeptidemolecule.基因是含有生物信息的DNA片段，根據(jù)這些生物信息可以編碼具有生物功能的產(chǎn)物，包括RNA和多肽鏈?；虻姆肿颖举|(zhì)

Genes:MolecularNature堿基配對(duì)A:TG:CTCGA雙鏈DNA的結(jié)構(gòu)右手方向雙股螺旋反向平行共同軸心大溝和小溝Bases

InDNA,therearefourheterocyclicbases:adenine(A)（腺嘌啉）andquinine(G)（鳥(niǎo)嘌啉）arepurines;cytosine(C)（胞嘧啶）andthymine(T)（胸腺嘧啶）arepyrimidines.InRNA,thymineisreplacedbythestructurallyverysimilarpyrimidine,uracil(U)（尿嘧啶）.Nucleosides

InRNAthesugarisribose（核糖）andthecompoundsareribonucleosides（核糖核苷）,orjustnucleosides,whereasinDNAitis2’-deoxyribose,andthenucleosidesarenamed2’-deoxyribonucleosides（2‘-脫氧核糖核苷）,orjustdeoxynucleosides.Base+sugar=nucleoside.Nucleotides

Inribonucleotides(核苷酸）(CMP,CTP,ATP,GTP,UTP),the2’-positionishydroxylgroup.Ifthesugarisdeoxyribose,thencompoundsaredeoxynucleotides（脫氧核苷酸）(dATP,dGTP,dCTP,dTTP;dNTP).

Base+sugar+phosphate=nucleotide.

PhosphodiesterbondsBetweenthe5’-positionofonesugarandthe3’-positionofthenext,forminga3’,5’-phosphodiesterbond.Nucleicacidsarehighlychargedpolymerswithanegativecharge（負(fù)電荷）oneachphosphate.帶負(fù)電荷的核酸在電場(chǎng)中向正極泳動(dòng)，泳動(dòng)速率與其分子量的對(duì)數(shù)近似負(fù)線性關(guān)系2000bp1500bp1000bp700bp500bp400bp300bp200bp100bp50bp900bp800bp700bp600bp500bp400bp300bpM12345Cathode(-)Anode(+)DirectionofmigrationSizeofDNAfragment1.5DNA/RNAsequenceThenucleicacidsequenceisthesequenceofbasesA,C,G,T/UintheDNAorRNAchain.Thesequenceisconventionallywrittenfromthefree5’-（游離5’-端）tothefree3’-end（游離3’-端)ofthemolecule.Examples:5’-ATAAGCTC-3’(DNA),5’-AUAGCUUGA-3’(RNA).DNAdoublehelixTwoseparateandantiparallelchains

（反向平行）ofDNAarewoundaroundeachotherinaright-handedhelical(coiled)path,withthesugar-phosphatebackbonesontheoutsideandthebases,pairedbyhydrogenbondingandstackedoneachother,ontheinside.Adeninepairswiththymine;guaninepairswithcytosine.Thetwochainsarecomplemently（互補(bǔ)）;onespecifiesthesequenceoftheother.Table1.1FormsofDNA

Form Pitch(nm) Residues Inclinationofbaseperturn pairfromhorizontal

A0.261120oB0.34100oC0.339.33D0.308.5Z0.45127o

RNA-DNAhybrid0.281120oRNAsecondarystructureMostRNAmoleculesoccurasasinglestrand（單鏈）,whichmaybefoldedintoacomplexconformation,involvinglocalregionsofintramolecularbasepairing（分子內(nèi)堿基配對(duì)）andotherhydrogenbondinginteractions.ModifiednucleicacidsCovalentmodificationsofnucleicacidshavespecificrolesinthecell.InDNA,thesearenormallyrestrictedtomethylationofadenineandcytosinebases,buttherangeofmodificationsofRNAismuchgreater.Example:methylation

（甲基化）oftheN-6positionofadenineandthe4-aminogroup,andthe5-positionofcytocine,phageDNAs;modificationsofRNA.ChemicalandPhysicalPropertiesofNucleicAcids核酸的物理化學(xué)性質(zhì)核酸的物理化學(xué)性質(zhì)Stabilityofnucleicacids（核酸穩(wěn)定性）Effectofacid（酸效應(yīng)）Effectofalkali（堿效應(yīng)）Chemicaldenaturation（化學(xué)變性）Viscosity（粘滯性）Stabilityofnucleicacids(核酸的穩(wěn)定性)AlthoughitmightseemobviousthatDNAdoublestrandsandRNAstructuresarestabilizedbyhydrogenbonding,thisisnotthecase.H-bondsdeterminethespecificityofthebasepairing,butthestabilityofanucleicacidhelixistheresultofhydrophobic（疏水）anddipole-dipoleinteractions（雙極-雙極相互作用）betweenthestacked（堆疊）basepairs.Effectofacid(酸效應(yīng))Highlyacidicconditionsmayhydrolyzenucleicacidstotheircomponents:bases,sugarandphosphate.Moderateacidcausesthehydrolysisofthepurinebaseglycosylicbonds

（糖苷鍵）toyieldapurinicacid（脫嘌啉核苷酸）.Morecomplexchemistryhasbeendevelopedtoremoveparticularbases,andisthebasisofchemicalDNAsequencing.Effectofalkali(堿效應(yīng))HighpHdenaturesDNAandRNAbyalteringthetautomeric（互變異構(gòu)）

stateofthebases

anddisruptingspecifichydrogenbonding.RNAisalsosusceptibletohydrolysisathighpH,byparticipationofthe2’-OHinintramolecularcleavageofthephosphodiesterbackbone.Chemicaldenaturation(化學(xué)變性)Somechemicals,suchasurea（尿素）andformamide（甲酰胺）,candenatureDNAandRNAatneutralpHbydisruptingthehydrophobicforcesbetweenthestacked（堆疊）bases.Viscosity(黏滯性)DNAsolutionshavehighviscosity（粘滯性）.LongDNAmoleculesaresusceptibletocleavagebyshearing（剪切）(orbysonication（超聲）insolution–thisprocesscanbeusedtogenerateDNAofaspecificaveragelength.SpectroscopicandThermalPropertiesofNucleicAcids

（核酸的熱力學(xué)和光吸收特性）UVabsorption（紫外吸收）Hypochromacity/hyperchromacity（減色/增色效應(yīng)）Quantitationofnucleicacids（核酸定量）PurityofDNA（DNA純度）Thermaldenaturation（熱變性）Renaturation（復(fù)性）Hybridization（雜交）UVabsorptionNucleicacidsabsorbUVlightduetotheconjugatedaromatic（共軛芳香基團(tuán)）natureofthebases;thesugar-phosphatebackbonedoesnotcontributeappreciablytoabsorption.ThewavelengthofmaximumabsorptionoflightbybothDNAandRNAis260nm(λmax=260nm).Theabsorptionpropertiesofnucleicacidscanbeusedfordetection（檢測(cè)）,quantitation（定量）andassessmentofpurity（純度估計(jì)）.3.2HypochromicityandHyperchromicityQuantitationofnucleicacidsAtaconcentrationof1mgml-1and1cmpathlength,double-strandedDNA（dsDNA）hasA260=20(or1OD260DNA=50μg/ml).RNAandsingle-strandedDNA（ssDNA）haveA260=25(or1OD260RNA/ssDNA=40μg/ml).ThevaluesforRNAandsingle-strandedDNAdependonbasecompositionandsecondarystructure.在波長(zhǎng)260nm處，1mg/ml濃度的DNA經(jīng)過(guò)1cm光路的吸光度值為20ODPuredsDNAhasanA260/A280of1.8,andpureRNAoneofaround2.0.Protein,withλmax=280nmhasaA260/A280ratioofless1(actuallyaround0.5).protein純度分析，PurityofDNAThermalDenaturation

熱變性IncreasedtemperaturecanbringaboutthedenaturationofDNAandRNA.RNAdenaturesgraduallyonheating,butdouble-strandedDNA‘melts’co-operativelytogivesinglestrandsatadefinedtemperature,Tm,whichisafunctionoftheG+CcontentoftheDNA.DenaturationmaybedetectedbythechangeinA260.Renaturation

復(fù)性DNArenaturesoncolding,butwillonlyformfullydouble-strandednativeDNAifthecoolingissufficientlyslowtoallowthecomplementaystrandsto

anneal（退火）.ThermaldenaturationandrenaturationHybridizationTherenaturationofregionsofcomplementaritybetweendifferentnucleicstrandsisknownashybridization.基因的功能

Genes:FunctionStoringInformationReplicationMutation遺傳信息的流向DNARNAProtein轉(zhuǎn)錄翻譯甲硫氨酸脯氨酸苯丙氨酸谷氨酸谷氨酸谷氨酰胺絲氨酸纈氨酸丙氨酸組氨酸丙氨酸甘氨酸谷氨酰胺DNARNA多肽鏈蛋白質(zhì)轉(zhuǎn)錄逆轉(zhuǎn)錄翻譯折疊組裝成熟復(fù)制中心法則Exon1Exon2Exon3Exon4Intron1Intron2Intron3

5’3’DNAPremRNAmRNATranscriptionmRNAprocessing3’-untranslatedregion5’-untranslatedregionTranslationProteinTherolesofRNA儲(chǔ)存遺傳信息(病毒，viruses)結(jié)構(gòu)分子(rRNAs)信息轉(zhuǎn)移(mRNAs)信息翻譯(tRNA)酶功能(核酶，ribozymes)TraditionalRNAsWhatisRNA?RibonucleicacidRibonucleotides(Ribose,base,&phosphate)TypesCoding:messengerRNA(mRNA)Non-coding:RibosomalRNA(rRNA)TransferRNA(tRNA)SmallnuclearRNA(snRNA)SmallnucleolarRNA(snoRNA)InterferenceRNA(RNAi)ShortinterferingRNA(siRNA)MicroRNA(miRNA)mRNAStructureCodingregionUntranslatedregions5’UTR7methyl-GcapBoundbycapbindingproteinsTranslationregulation3’UTRStabilityelementsSubcellularlocalization(zipcodes)poly(A)tailmRNA基因的命名

GeneNomenclature基因命名的基本原則人類(lèi)基因命名委員會(huì)(HumanGeneNomenclatureCommittee)的基本指導(dǎo)方針：應(yīng)隨新技術(shù)的發(fā)展而發(fā)展，而不應(yīng)受到歷史的局限性?；蛎幕驹瓌t：簡(jiǎn)明、獨(dú)特、能夠表達(dá)基因的特征或功能。基因符號(hào)的使用規(guī)則：獨(dú)特、簡(jiǎn)短、僅用拉丁字母和阿拉伯?dāng)?shù)字，不應(yīng)含有標(biāo)點(diǎn)符號(hào)。GeneNomenclature基因命名：瀏覽倫敦大學(xué)學(xué)院網(wǎng)站http://www.gene.ucl.ac.uk/nomenclature/命名指南：瀏覽倫敦大學(xué)學(xué)院網(wǎng)站http://www.gene.ucl.ac.uk/nomenclature/guidelines.html人類(lèi)基因組組織(HUGO)/Guidelinesforhumangenenomenclaturewerefirstpublishedin1979[1],whentheHumanGeneNomenclatureCommitteewasfirstgiventheauthoritytoapproveandimplementhumangenenamesandsymbols.Updatesoftheseguidelineswerepublishedin1987[2],1995[3],and1997[4].

Withtherecentpublicationsofthecompletehumangenomesequencethereisanestimatedtotalof26,000-40,000genes,assuggestedbytheInternationalHumanGenomeSequencingConsortium[5]andVenteretal.[6].

Thus,theguidelineshavebeenupdatedtoaccommodatetheirapplicationtothiswealthofinformation,althoughsymbolsarestillonlyassignedwhenrequiredforcommunication.TheseupdateswerederivedwithinputfromtheHUGOGeneNomenclatureCommittee(HGNC)InternationalAdvisoryCommitteeandattendeesoftheASHG01NWGeneNomenclatureWorkshop.

AllapprovedhumangenesymbolscanbefoundintheGenewdatabase[7]./cgi/content/full/30/1/1691.Criteriaforsymbol