GMan - Polymerases

  1. RNA Polymerase
    ssDNA Template
  2. DNA Polymerase
    • dsDNA Template
    • Can Only extend
  3. Primase
    • E.coli enzyme
    • Builds RNA primer from scratch
  4. RNA Primer length
  5. Simple overview of DNA polymerase mechanism
    • DNA polymerases uses Mg2+ to attach a dNTP to the 3' end of the nascent strand
    • A pyrophosphate is released
  6. Phosphodiester bond
    • Links the 5' phosphate to the 3' O
    • Formed by a condensation reaction
  7. Who isolated RNA Pol I
    Arthur Kornberg
  8. Who isolated RNA Pol III
    Thomas Kornberg - Also discovered Pol II
  9. RNA Pol I
    • Most Abundant
    • Relatively minor role
    • Primer removal, DNA repair
  10. RNA Pol III
    • Regarded as the most active
    • Chromosome replication
  11. DNA Pol IV, V uses
    SOS Repair
  12. All three DNA polymerases can elongate DNA? (T/F)
    T - But they can't all initiate it
  13. Exonuclease Activity
    • Proofreading
    • Direction, 3'->5'
    • DNA Pol I, II and III
  14. 5' -> 3' Exonuclease activity
    • DNA Pol I
    • Primer removal
  15. Fidelity
    • How accurate a polymerase is
    • Includes low misincorporation (errors) and proofreading
  16. Processivity
    The number of nucleotides a polymerase can churn out before dropping off
  17. Highest fidelity polymerase
    DNA Pol III
  18. Highest and lowest processivity polymerase
    • Pol III - 500k
    • Pol I - 200
  19. Effect of proofreading on fidelity
    Can increase fidelity by two orders of magnitude
  20. Names of the DNA Pol III subunits
    • Primer Breakdown
    • Replace gap from the primer with DNA
  21. Rate of DNA Pol III
    1000 nt/sec
  22. Subunits of RNA Pol III found in triplet
    α, ε, θ
  23. Subunits of the DNA Pol III γ complex
    γ, δ, δ', χ, ψ
  24. Other subunits of DNA Pol III
    β, τ
  25. DNA Pol III holoenzyme
    • Composed of two enzymes
    • DNA Pol III
    • Sliding clamp loader
  26. Role of the τ proteins in DNA Pol III
    Hold the DNA Pol III core proteins together
  27. Replisome
    DNA Pol III Holoenzyme + Helicase + Primase
  28. Lagging strand synthesis uses...
    The second arm of DNA Pol III
  29. Eukaryotic version of the sliding clamp
    Proliferating cell nuclear antigen (PCNA)
  30. Primosome
    Heliacase + Primase at replication fork
  31. How long are okazaki fragments?
    1000nt - 2000nt
  32. DNA ligase
    Fixes the gap between okazaki fragments
  33. DNA ligase mechanism
    • A lys (NH2) on the catalytic site binds AMP (ATP -> AMP, PPi released)
    • The ligase formas a phosphoanhydride bond with the 5' phosphate adjacent to the nick
    • The 3' OH then attacks the phosphoanhydride
  34. The DNA moves, but the replisome is stationary. (T/F)
  35. Tus proteins
    • Terminator utillization substance
    • Stop the replication fork moving
    • Only act in one direction
Card Set
GMan - Polymerases