Biochem 501: Part IV.6: Transcription

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Biochem 501: Part IV.6: Transcription
2013-12-06 02:21:08
Biochem 501 FINAL

Biochem 501 FINAL
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  1. Transcription of RNA
    • RNA polymerase binds to DNA promoter
    • Polymerase synthesizes RNA based on DNA sequence
    • DNA sequence signals the ends of transcription
  2. An RNA sequence is same as the:
    DNA Non-template "coding" strand
  3. RNA transcription uses how many strands of DNA
  4. RNA can initiate RNA synthesis ___ and starts at a ___
    de novo, promoter
  5. E. coli sigma 70 holoenzyme
    • Well studied RNA polymerase
    • Composed of core enzyme with additional subunit called sigma, which provides initiation specificity. Core enzyme does elongation, sigma does initiate. Sigma subunits can vary
  6. How many types of Eukaryotic RNA polymerase
  7. Experimental approaches that identified promoters
    • Genetics: mutations that change the
    • frequency of transcription initiation

    • Biochemistry: DNA sequences that initiate
    • transcription by RNA polymerase

    Sequence comparisons: common DNA sequences found near RNA 5’ end

    Footprint assays: DNA sequences bound by RNA polymerase
  8. Typical Eukaryotic promoter
    TATA box
  9. Prokaryotic RNA synthesis
    • RNA polymerase binds DNA promoter
    • Local denaturation at promoter
    • RNA primer synthesis
    • sigma unit released, RNA pol moves off promoter
    • Polymerase synthesizes RNA based on DNA synthesis
    • DNA sequence signals end of RNA transcription
  10. Steps in transcription initiation
    • Closed complex formation: RNA polymerase (R) binds to double stranded DNA promoter (P). R+P becomes RPc.
    • Open complex formation: RPc isomerizes to form open complex in which there is a localized denaturation of DNA. Template DNA located in active site of enzyme (RPc to RPo)
    • Initiation: Ribonucleoside triphosphates condensed to form short oligonucleotides complementary to template strand. (RPo + rNTPs yields short oligonucleotides +RPi)
    • Promoter clearance: sigma subunit and promoter released. Core R starts extending oligonucleotides to long RNA molecule.
  11. How does RNA polymerase add nucleotides
  12. Termination of transcription
    • Intrinsic Termination (Rho-independent): A G-C rich RNA sequence forms hairpin structure, followed by UUUU which destabilizes RNA polymerase nucleoprotein complex, termination
    • Rho-stimulated termination: Bacterial protein called rho, or p, terminates transcription at specific regions. Mechanism unclear.
  13. Eukaryotic RNA polymerases
    • Pol I, II, III
    • Pol I, III: code for non-coding RNA (I: rRNA, III: tRNA, some rRNA, additonal RNAs)
    • Pol II: Codes for mRNA--->ribosome--->protein
  14. Eukaryotic Transcriptional Initiation requires:
    Multiple Protein Components
  15. Initiation factors associate dynamically with the promoter
  16. Rifampin does what
    • Blocks bacterial RNA polymerase, prevents release of sigma subunit and subsequent elongation
    • Tuberculosis