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Chapter 21 (part 1)
Transcription
Central Dogma
RNA Content of E. coli Cells
typeSteady State Levels
Synthetic Capacity
Stability
rRNA 83% 58% High
tRNA 14% 10% High
mRNA 3% 32% Very Low
E. Coli RNA Polymerase• RNA polymerase core
enzyme is a multimeric protein ’
• The ’ subunit is involved in DNA binding
• The subunit contains the polymerase active site
• The subunit acts as scaffold on which the other subunits assemble.
• Also requires -factor for initiation –forms holo enzyme complex
Site of DNA binding and
RNA polymerization
General Gene Structure5’ 3’Transcribed region terminatorPromoter
Gene Promoters
Other -Factors• Standard genes – 70
• Nitrogen regulated genes – 54
• Heat shock regulated genes – 32
Transcriptional Initiation
Closed complex
Open complex
Primer formation
Disassociation of -factor
Pausing induces termination
3’end tends to be AU rich easily to disrupt during pausing. Leads to disassembly of RNA polymerase complex
Rho Dependent Termination
• rho is an ATP-dependent helicase
• it moves along RNA transcript, finds the "bubble", unwinds it and releases RNA chain
Eukaryotic RNA Polymerases
type Location ProductsRNA polymerase
I Nucleolus rRNA
RNA polymerase II
Nucleoplasm mRNA
RNA polymerase III
Nucleoplasm
rRNA, tRNA, others
Mitochondrial RNA polymerase
Mitochondria
Mitochondrial gene
transcripts
Chloroplast RNA polymerase
Chloroplast
Chloroplast gene
transcripts
Eukaryotic RNA Polymerases
• RNA polymerase I, II, and III
• All 3 are big, multimeric proteins (500-700 kD)
• All have 2 large subunits with sequences similar to and ' in E.coli RNA polymerase, so catalytic site may be conserved
Eukaryotic Gene Promoters• Contain AT rich concensus sequence
located –19 to –27 bp from transcription start (TATA box)
• Site where RNA polymerase II binds
Transcription Factors• TFAIIA, TFAIIB –
components of RNA polymerase II holo-enzyme complex
• TFIID – Initiation factor, contains TATA binding protein (TBP) subunit. TATA box recognition.
• TFIIF – (RAP30/74) decrease affinity to non-promoter DNA