K原核生物的转录

1、1会计学K原核生物的转录原核生物的转录(-) strand is antisense strand. (+) strand is sense strandATACGTATGC+1promoterterminatorTranscribed regionRNADNATranscriptionAntisense strandAUACGStructure of a transcription unitWhat is a promoter (启动子启动子)?The sequence of DNA needed for RNA polymerase to bind to the template and

2、accomplish the initiation reaction 位于蛋白质编码区的上游（位于蛋白质编码区的上游（5端）端）含有短而保守的含有短而保守的DNA序列，不同基因的启序列，不同基因的启动子中序列经常是保守的。动子中序列经常是保守的。Transcription bubbleElongationRNA hairpin (发卡发卡) structure5 NNNNAAGCGCCGNNNNCCGGCGCUUUUUU -OHNN NN CGCCGCGGCCGGCAUAU NNNN UUUU-OHSteps for RNA transcription2. Requires DNA for a

3、ctivity and is most active with a double-stranded DNA as template. 5 3 synthesis(NMP)n + NTP (NMP)n+1 + PPiRNA polymerase: synthesis of RNA strand from DNA template.1. Requires no primer for polymerization3. Require Mg2+ for RNA synthesis activity4. All RNA polymerases lack 3 5 exonuclease activity,

4、 and one error usually occurs when 104 to 105 nucleotides are incorporated. 5. usually are multisubunit enzyme, but not always. 6. Different from organism to organism7. E. coli has a single DNA-directed RNA polymerase that synthesizes all types of RNA. Both 起始起始 & 延伸延伸起始起始 only36.5 KD36.5 KD151

5、KD155 KD11 KD70 KD465kd RNA synthesis rate: 40 nt per second at 37oC 非球形结构，圆柱形孔道旁有一突起，非球形结构，圆柱形孔道旁有一突起，孔道大小表明孔道大小表明the enzyme complex can bind directly to 16 bp of DNA. The whole polymerase binds over 60 bp.E. coli RNA polymerase:全酶全酶: 2 for initiation核心酶核心酶: 2 for elongationRNA pol 亚基亚基E. coli poly

6、merase: subunit E. coli polymerase: subunit E. coli polymerase: factor E. coli polymerase: factor K3 The E. coli 70 promoter1. Promoter sequences: -10 sequence and -35 sequence2.Transcription start site3.Promoter efficiencyATACGTATGC+1promoterterminatorTranscribed regionRNADNATranscriptionAntisense

7、strandDifferent promoters result in differing efficiencies of transcription initiation, which in turn regulate transcription. AUACG-5-8 bp- GC T ATTGACATATAAT-16-18 bp-+1-35 sequence-10 sequence A purine in 90% of all genes G is more common at position +1 than A Often, 起始位点两侧为C and T (i.e. CGT or CA

8、T)The sequence around the start site influences initiationThe sequences of five E. coli promotersConsensus TTGACATATAATDNA unwinding RNA chain initiation RNA chain elongation RNA chain termination Rho-dependent termination BindingUnwindingInitiationElongationTerminationantisenseRNA chain 终止Terminati

9、on: dissociation of RNA re-annealing of DNA release of RNA polTerminator sequence 终止序列 (stop signal 停止信号): RNA hairpin very commonAccessory rho proteinA model for -independent termination of transcription in E. coli. The A-U base-pairing is less stable that favors the dissociation2. Rho-dependent Te

10、rmination. lNo U stretch at the 3 end of the RNAlRho protein (hexameric protein) binds to certain RNA structure (72bp) moves along the nascent RNA toward RNA pol complex stop at the rho-dependent transcription terminator. RNA polDNA polTemplatedsDNA is betterss/dsDNARequire primerNoYesInitiationprom

11、oteroriginelongation40 nt/ sec900 bp/secterminatorSynthesized RNA Template DNASummary:1. General features of RNA pol.2. E. coli RNA pol: holoenzyme ( 2 ) and core enzyme ( 2 ). Role of sigma factor.3. E. coli 70 promoter: -35 sequence (TTGACA), -10 (TATAAT), & +1 (purine, G A)4. Transcription: initiation, elongation and termination (hairpin and rho factor)ATACGTATGC+1promot