Mol Bio Mid Review part 5

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  1. The __ of the primer attacks the __ of the entering nucleotide and makes an ester bond with it. One __ is released from each added nucleotide and then is __
    • 3'-OH;
    • phosphate;
    • pyrophosphate;
    • hydrolyzed to 2 phosphates
  2. __ are discriminated by the steric exclusion. There is a __ in the nucleotide binding pocket of the active site
    • NTPs (building blocks of RNA);
    • discriminator amino acid
  3. steric constrict:
    • 2`-OH of NTPs + discriminator aa → displacement of alpha phosphate → no bond with the 3`-OH (of the extending strand)
    • Image Upload
  4. palm domain of DNA polymerase
    • the catalytic site
    • involves binding of metal ions (Mg 2+ or Zn 2+)
    • monitoring base pairing of the recently added nucleotide
  5. palm domain: One metal ion __. The 2nd metal ion __
    • reduces affinity of 3'OH for its H → making 3'-0- → attacking alpha phosphates
    • coordinates with the negative charges of beta and gamma phosphates of the entered dNTP (nucleotides) and stabilizes the released pyrophosphates
  6. fingers domain of DNA polymerase
    • important for catalysis
    • Several amino acid residues of "fingers" make bonds with the incoming dNTP (nucleotides)
  7. If base pairing is correct, the O-helix of fingers __
    moves by 40 degrees → closing the hand (enclosing dNTP) → making stacking bonds with the nucleotide → pushing it to the proximity of the metal ions of the enzyme → stimulating catalysis
  8. Before connecting the Okazaki fragments, the primers are removed by __ and then replaced by DNA. The last nucleotide of the primer attached to the DNA is removed by the __.  A __ fills the generated gap. The last nucleotide added to the gap is joined to the next nucleotides by a phosphodiester bond generated by the enzyme __
    • RNase H;
    • 5' exonuclease (because RNase H cannot remove it);
    • DNA polymerase;
    • DNA ligase
    • Image Upload
  9. Hexameric ring shape DNA helicase __
    opens the 2 strands from each other. There is a pore in the center of this ring that ssDNA passes through
  10. Each subunit of helicase has a hair pin like structure that __
    binds to a phosphate and its 2 adjacent deoxyriboses

    (These hair pins are arranged like a right handed staircase, each one attached to one phosphate and the 2 deoxyriboses.)
  11. The DNA helicase structure uses ATP to __
    pull ssDNA through the pore one nucleotide after the other
  12. DNA helicase has a polarity (moving 5' → 3' or 3' → 5') depending on the strand the helicase is attached to. For the lagging strand, it's __
    5' → 3'
  13. Unwinding of DNA during replication → sending turns down the DNA → making __ supercoil

    Supercoil must be removed otherwise the replication cannot continue.
  14. During replication, the supercoil is removed by __

    Topoisomerase I and II break one or both strands and pass the other strand (Topo I) or both strands (Topo II) and relieve the supercoil.
  15. Processivity is increased by the association of DNA pol with an assembly of proteins known as __
    DNA sliding clamp

    • Sliding clamp is ringed around the DNA and slides along the DNA without getting dissociated
    • Sliding clamp also binds to DNA pol bound to the primer:template junction.
    • The whole complex moves along the DNA
    • Image Upload
  16. If the DNA pol is detached from the DNA the clamp keeps it around, __
    DNA pol reattaches and restarts the polymerization → increased processivity
  17. In E. coli, __ is made of 3 copies of DNA pol III and one (5 unit) sliding clamp loader.
    DNA pol III holoenzyme
  18. The holoenzyme interacts with DNA helicase by the t proteins. After this interaction, __
    the helicase activity increases by 10 fold. Helicase generates the ssDNA with the same rate as DNA polymerization
  19. Helicase also interacts (weakly) with the primase, __
    enhancing activity of primase by 1000 fold
  20. All assembled proteins together at the replication fork are also known as __
  21. DnaB begins to increase the single-stranded region within the origin, enabling the enzymes involved in the elongation of DNA replication to attach. DNA polymerase is attached to the primer:template junction. →
    • start of DNA replication. (attachment of 3 sliding clamp, fork is completed). This represent the __ of replication of E.coli
    • end of the initiation phase
  22. steps in the initiation phase of DNA replication
    • 1. binding of origin recognition complex (ORC) to the origins of replication on the DNA (happens in all phases of the cell cycle)
    • 2. formation of preRC (rereplicative complex): occurs in early G1 phase. At this step, helicases are inactive and do not unwind the double strand. Helicase loading proteins are released.
    • 3. Activation of helicases (occurs in S phase) → unwinding the double helix and loading of DNA replication enzymes onto the 2 single-stranded templates (completion of replisome)
  23. DNA replication happens once per cycle because:
    • There is no formation of new preRCs during S phase
    • In the S phase, only the previously made preRCs are becoming activated by the activity of regulatory proteins of the S phase
    • After mitosis (early G1), new preRCs are made again, which in turn become activated in the next S phase
  24. As S phase is about to start, regulating proteins of the S phase phosphorylate and thereby inhibit the ORC and Cdc6 proteins, __ before the mitosis is completed.
    preventing formation of new preRCs
  25. After mitosis, regulating proteins of the S phase are __
    downregulated → Cdc6 and Cdt1 (that help to load helicase) are activated → new round of loading the helicases
Card Set:
Mol Bio Mid Review part 5
2017-02-14 02:59:34
bio replication

Week 6
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