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110C LEC 8 RNA structure and synthesis

110C LEC 8 RNA structure and synthesis - BCH 110C LECTURE 8...

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BCH 110C LECTURE 8 RNA structure and synthesis Lodish 6 th ed. Pages 11-12, 17-18, 24-25, 111-112; Chapters 4.1-4.2; 7.2
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Central Dogma of Molecular Biology General Pattern of Information Flow in Cells RNA Informational : mRNA Component of protein synthetic apparatus : rRNA Catalytic/functional : e.g. self splicing introns, ribozymes, telomerase, ribonuclear proteins, peptidyltransferase large rRNA. Regulatory RNA : miRNA, siRNA, piRNA, rasiRNA Genomic : RNA viruses (HIV)
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RNA Structure
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RNA is normally single stranded but forms intrastrand H-bonds Secondary structure of 16S rRNA from E. coli RNA usually has extensive secondary structure and tertiary
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RNA is synthesized 5’ to 3’ •H-bonding between the base of an incoming nucleotide triphosphate and the template DNA strand. •Nucleophilic attack of the 3’OH on the α -phosphate of the incoming nucleotide triphosphate. •A new phosphodiester bond is formed. •Pyrophosphate is the leaving group. No primer is required
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E. Coli RNA Polymerase ( α 2 ββ ωσ ) One σ subunit per polymerase Core polymerase Holoenzyme Subunit Number kDal Function 2 36.5 - ß 1 151 Nucleotide binding; Associates with DNA Start Site ß’ 1 155 Polymerization; Associates with DNA Start Site 1 11 - 70 1 70 Positions polymerase for initiation on most promoters 54 - 54 Positions polymerase for initiation on promoters for nitrogen starvation genes
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Not for memorization
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E. Coli RNA Polymerase Core enzyme: 2  ω – NOT capable of promoter recognition • Holoenzyme: 2  ωσ – Capable of promoter recognition – The σ bound to core polymerase determines which promoters are transcribed by the polymerase
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σ 70 Promoter Recognition by E. coli RNA Polymerase By Convention +1 + - General plan: TGTTGACAT---11-15 bp----TATAAT----5-8 bp----Initiation Site Pribnow or TATA box -35 region σ recognizes both TATA & -35 region
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Promoter Strength Bacteria have 1000-4000 genes Every promoter has a slightly different sequence These variations make some promoters better at facilitating initiation (“stronger”) and some worse (“weaker”) Generally, the closer a promoter is to the consensus sequence the stronger the promoter
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Initiation by E. coli RNA Polymerase 1. -Holoenzyme forms -Holoenzyme binds DNA and slides along it in a series of binding and release steps until a promoter is encountered -It binds the promoter in a “closed complex” 2. In an energy requiring process, polymerase unwinds the DNA, and rearranges into an “open complex” 3. Transcription begins -Initial synthesis inefficient 4. After 10 nucleotides σ dissociates -RNA placed in exit channel
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