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Understanding E. coli RNA Polymerase and Eukaryotic Transcription Mechanisms

This chapter explores the roles and structures of the RNA polymerase enzyme in E. coli, highlighting its multimeric composition and the importance of each subunit in DNA binding and RNA synthesis. The initiation process, involving s-factors and promoter regions, is detailed alongside transcriptional dynamics, including pausing and termination mechanisms like rho-dependent termination. Additionally, the chapter discusses eukaryotic RNA polymerases I, II, and III, their similarities to E. coli polymerases, and the role of transcription factors in eukaryotic gene promoters, particularly the significance of the TATA box in transcription initiation.

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Understanding E. coli RNA Polymerase and Eukaryotic Transcription Mechanisms

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Presentation Transcript

  1. Chapter 21 (part 1) Transcription

  2. Central Dogma

  3. RNA Content of E. coli Cells

  4. E. Coli RNA Polymerase • RNA polymerase core enzyme is a multimeric protein a2,b, b’, w • The b’ subunit is involved in DNA binding • The b subunit contains the polymerase active site • The a subunit acts as scaffold on which the other subunits assemble. • Also requires s-factor for initiation –forms holo enzyme complex Site of DNA binding and RNA polymerization

  5. Promoter Transcribed region terminator 5’ 3’ General Gene Structure

  6. Gene Promoters

  7. Other s-Factors • Standard genes – s70 • Nitrogen regulated genes – s54 • Heat shock regulated genes – s32

  8. Transcriptional Initiation Closed complex Open complex Primer formation Disassociation of s-factor

  9. Pausing induces termination 3’end tends to be AU rich easily to disrupt during pausing. Leads to disassembly of RNA polymerase complex

  10. Rho Dependent Termination • rho is an ATP-dependent helicase • it moves along RNA transcript, finds the "bubble", unwinds it and releases RNA chain

  11. Eukaryotic RNA Polymerases

  12. 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

  13. Eukaryotic Gene Promoters • Contain AT rich concensus sequence located –19 to –27 bp from transcription start (TATA box) • Site where RNA polymerase II binds

  14. 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

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