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Ann's M2M Cards - weeks 1, 2 > Replication (Huang) > Flashcards

Replication (Huang) Flashcards

1
Q

DNA replication is bi or uni - directional

A

Bidirectional

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2
Q

How many origins in prokaryotes?

A

One

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3
Q

How many origins in eukaryotes?

A

Hundreds

some used more than others

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4
Q

Define semiconservative

A

Each new strand is one old strand and one new strand.

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5
Q

Replication fork

A

Site where replication is occurring

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6
Q

What recognizes and binds to origin?

A

Origin binding proteins

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7
Q

Where are origins likely to be?

A

AT rich sequences

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8
Q

What unwinds the parental strands?

A

Helicase

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9
Q

What prevents super coiling?

A

Topoisomerases in eukaryotes

Gyrase in prokaryotes

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10
Q

What inhibits dna gyrase?

A

quinolones

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11
Q

what are quinolones?

A

class of broad spectrum antibiotics

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12
Q

Direction of DNA replication?

A

5’ —> 3’

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13
Q

What catalyzes the synthesis of DNA?

A

DNA polymerase

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14
Q

How does DNA polymerase work?

A

adds deoxyribonucleotides to the 3’- OH of RNA primers then to 3’- OH of growing DNA strands

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15
Q

How many DNA polymerases in prokaryotic DNA replication?

A

2

Pol I and Poll III

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16
Q

Which is used more? Pol I or Pol III

A

Poll III

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17
Q

Why is Poll III used more in prok. DNA repl

A

has a sliding clamp so can work over a longer distance (more processive manner)

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18
Q

What does Pol I do?

A

Clean-up work during DNA replication and repair

Replaces RNA primers via
5’ - 3’ exonuclease activity
5’ - 3’ DNA polymerase activity

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19
Q

How many polymerases in Eukaryotic DNA replication?

A

3
Pol alpha
Pol delta
Pol epsilon

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20
Q

What does Pol alpha do?

A

is a holoenzyme - multi-protein complex

Primase activity
DNA polymerase activity

Synth. first ~20 residues after RNA primer

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21
Q

Does Pol alpha have proofreading activity?

A

No

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22
Q

What does Pol delta do?

A

Synthesis of lagging strand

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23
Q

What does Pol epsilon do?

A

Synthesis of leading strand

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24
Q

Can DNA replication start ‘de novo’?

A

No. Requires an RNA primer

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25
Q

What prepares the RNA primer?

A

primase

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26
Q

What does SBB do?

A

Single Strand Binding protein holds and protects the single strands unwound by helicase during replication

27
Q

What does helicase do?

A

unwinds parent DNA strand

28
Q

What does origin binding protein do?

A

recognizes and binds to origin

29
Q

What serves as primer for DNA synth?

A

RNA

30
Q

How long is the primer?

A

~10 residues of RNA

31
Q

What synthesizes the first ~20 DNA residues onto the primer?

A

Pol alpha

32
Q

What synthesizes the leading strand in eukaryotes?

A

Pol epsilon

33
Q

What synthesizes the lagging strand in eukaryotes?

A

Pol delta

34
Q

What does primase do?

A

synth the RNA primer

35
Q

How do the RNA primers get chosen?

A

use the DNA template

36
Q

Replication fork?

A

place where DNA replication is ocurring simultaneously on both parent strands but in opp. directions

e.g. both grow in 5’ - 3’ direction along the growing strand but spatially opposite direction

37
Q

Which strand grows towards replication fork?

A

leading strand

38
Q

Leading strand grows _______________.

A

continuously

39
Q

Lagging strand grows _____________.

A

discontinuously. e.g. okazaki fragments

40
Q

Which strand grows away from the replication fork?

A

lagging strand

41
Q

What are okazaki fragments?

A

short fragments of DNA formed in synth of lagging strand

42
Q

how long are okazaki fragments?

A

Euk: 100-200 bp
Prok: 1000 - 2000 bp

43
Q

How are RNA primers removed?

A

5’ - 3’ exonuclease (DNA pol I in e.coli)

44
Q

How are gaps from RNA primers filled?

A

with appropriate DNA using template (by pol I in e.coli)

45
Q

In E. Coli what are the functions of Pol I?

A
  1. 5’ - 3’ DNA polymerase activity requiring 3’-OH primer
  2. 5’ - 3’ exonuclease activity to remove RNA primer
  3. 3’ - 5’ exonuclease activity for proofreading
46
Q

How are okazaki fragments joined?

A

by DNA ligase

47
Q

What does DNA ligase do?

A

forms phosphodiester bonds between 3’-OH and 5’-phosphate of two polynucleotide chains to join okazaki fragments

48
Q

Overall fidelity of DNA replication?

A

error rate of 10^-9 to 10^-10

49
Q

2 ways polymerases discriminate between correct and incorrect nucleotide?

A
  1. Hydrogen bonding between complimentary nucleotides (A-T, C-G)
  2. Geometry of the A-T and C-G base pairs that allow them to fit into the active site of polymerase
50
Q

H-bonding and bp geometry account for what level of accuracy?

A

1 error in 10,000 to 100,000 correct nucleotieds

51
Q

If an error occurs, how is it fixed during replication?

A

Proofreading during replication by 3’-5’ exonuclease activity associated with polymerase complex

52
Q

Proofreading increases the accuracy of replication by 100 - 1,000 fold. What is the final error rate?

A

1 error per million to 100 million correct nucleotides

53
Q

How else can DNA repair occur other than during replication?

A

Post-replicational repair…see DNA repair lecture

54
Q

What is synthesis of DNA from RNA?

A

reverse transcription

55
Q

Name two instances where reverse transcription takes place?

A
  1. Retroviruses

2. Telomerase activity

56
Q

What does telomerase do?

A

has reverse transcriptase activity
carries its own RNA template
restores ends of chromosome (telomeres) in human cancer and stem cells

57
Q

What is the end replication problem?

A

leading strand can be synthesized to the very end but the lagging strand cannot

58
Q

Why can the lagging strand NOT be synthesized to the very end?

A

Need an RNA primer ot begin synth. of each piece.

The last piece has no attachment for primer

59
Q

What is result of lagging strand not going to the very end of replication?

A

Telomere gets shorter with each round of replication.

60
Q

In cancer cells is telomerease turned on or off?

A

On. (de-repressed) Bad.

blocks normal cell death promoting tumor growth.

61
Q

In normal cells is telomerase on or off?

A

Off (repressed)

62
Q

What is added to the CTD of poll II and what is its function?

A

The CTD of pol II is phosphorylated. This attaches to the 5’-cap on the growing RNA strand and serves as a “landing pad” for the incoming nucleotides

63
Q

What protein is a trimer of the PCNA protein and what is its function?

A

The DNA “sliding clamp”
holds DNA template in the polymerase
aids in processivity

64
Q

What is the ORC?

A

Origin recognition complex

Ann's M2M Cards - weeks 1, 2 (16 decks)

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