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Biochemistry > L17-18 DNA Replication > Flashcards

L17-18 DNA Replication Flashcards

1
Q

Meselson & Stahl (1958)

A
  • DNA is synthesised in a heavy medium (15 N)
  • DNA is then moved to the light medium (14 N) and replicated one cycle
  • This is then repeated for 2, 3 cycles etc.
  • Density of DNA is measured after each stage using density-gradient equilibrium sedimentation in CsCl
  • This showed an intermediate weight DNA after replicating the the light medium once
  • This supported semi-conservative replication
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2
Q

DNA polymerase was discovered by…

A

Korburg (1958)

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3
Q

DNA polymerase adds…

A

dNTP precursors to DNA

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4
Q

DNA polymerase is a template…

A

Directed enzyme

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5
Q

DNA polymerase ( does / does not ) require a primer

A

Does

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6
Q

The template strand catalyses the formation of phosphodiester linkages if…

A

The bases are complimentary

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7
Q

DNA polymerase catalyses…

A

Nucleophilic attack on the inner phosphorus of the new molecule by the 3’ hydroxyl of the previous DNA molecule

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8
Q

DNA polymerase moves from ( 3’ / 5’ ) to ( 3’ / 5’)

A

5’ to 3’ (relative to the new strand)

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9
Q

DNA polymerase can remove mismatched nucleotides as it has…

A

3’ to 5’ exonuclease activity

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10
Q

DNA replication in E.Coli has…

1) two origin points
2) three origin points
3) one origin point
4) many (>3) origin points

A

3) one origin point

Known as oriC

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11
Q

OriC contains 5 copies of…

A

dnaA binding sequence

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12
Q

The dnaA hexamer binds to the sequences at oriC, which causes (2)…

A
  • Strain on the DNA backbone

* unwinding at the weak AT rich array upstream of OriC

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13
Q

dnaB helicase is recruited by…

A

dnaA

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14
Q

dnaB helicase is loaded around ( ssDNA / dsDNA ) at the ( 3’ / 5’ ) end

A

ssDNA at the 5’ end

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15
Q

ATPase-dependent translocation is carried out by…

A

dnaB helicase

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16
Q

The model of dnaB as loading around one strand of the DNA is known as the…

A

Strand exclusion model

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17
Q

Exposed single stranded DNA is stabilised by the binding of…

A

SSBs (single-stranded binding proteins)

18
Q

SSBs are

a) monomeric
b) trimeric
c) tetrameric
d) hexameric

A

c) tetrameric

They are made up of four monomers

19
Q

Trying to unwind a helix can cause…

A

Supercoiling of the remaining helix!

20
Q

To relax the dsDNA and prevent supercoiling…

A

Topoisomerase I is recruited

21
Q

Topoisomerase II prevents tangling and…

A

Catalyses the untangling of DNA duplexes by allowing two strands to pass through one another

22
Q

Topoisomerase I relaxes the DNA by…

A

1 turn

23
Q

DNA replication is:

a) initiated without the need for a primer
b) initiated by a DNA primer
c) initiated by an RNA primer
d) initiated by either a DNA or RNA primer

A

c) initiated by an RNA primer

This might be evidence for the RNA world hypothesis

24
Q

Primase can:

a) initiate but not extend
b) extend but not initiate
c) initiate and extend
d) none of the above

A

a) initiate but not extend

25
Q

Conversely, DNA polymerase can:

a) initiate but not extend
b) extend but not initiate
c) initiate and extend
d) neither extend nor initiate

A

b) extend but not initiate

Hence why a the primer is needed

26
Q

Replication of the leading strand is catalysed by DNA polymerase ( I / II / III / IV )

A

DNA polymerase III

DNA polymerases I & II are involved with lagging strand synthesis

27
Q

The DNA polymerase III core is made up of the (3)…

A

α, ε and θ subunits

alpha, epsilon and theta

28
Q

The α-subunit of DNA polymerase III catalyses…

A

Base addition and maintains the correct geometry for successful base pairing!

29
Q

The error rate of DNA polymerase III is reduced from 1 in 10^3 to around 1 in 10^6 by the…

A

Exonuclease which performs a proofreading function, cutting off incorrect base pairs and sending the strand back

30
Q

The β2 subunit is the…

A

Sliding clamp

31
Q

The β2 subunit increases the processivity of the enzyme, resulting in a rate of…

A

1000 nucleotides per second!

32
Q

Unlike dnaB helicase, the hole in the similarly donut-shaped β2 subunit is…

a) larger (35Å in diameter) allowing dsDNA through
b) smaler (35Å in diameter) not allowing dsDNA through
c) larger (70Å in diameter) allowing ssDNA through

A

a) larger (35Å in diameter) allowing dsDNA through

33
Q

The lagging strand requires…

a) a primer at the start
b) a primer for each fragment
c) a primer every 40-50 nucleotides

A

b) a primer for each fragment

34
Q

A significant amount of newly synthesised DNA exists as small pieces, known as…

A

Okazaki fragments

35
Q

DNA polymerase can only synthesise in the…

A

5’ to 3’ direction

36
Q

DNA polymerase I…

A

removes RNA primers and synthesises DNA

37
Q

DNA polymerase I is ( faster / slower / the same speed) as DNA polymerase III

A

Slower at about 10nt/s versus 1000nt/s

38
Q

DNA ligase fills in the…

A

‘nicks’ between okazaki fragments, catalysing phosphodiester bond formation

39
Q

DNA polymerase I has ( 5’ → 3’ / 3’ → 5’ ) exonuclease activity

A

5’ → 3’

40
Q

The trombone model describes the… 🎺

A

Lengthening and shortening of the visible ssDNA loop of lagging strand created by DNA polymerase III holoenzyme during the replication process

41
Q

There are an array of DNA repair proteins which are at work…

A

Constantly! 👷‍♀️

42
Q

Dideoxy Sequencing is a method of DNA sequencing pioneered by…

A

Sanger (1977)

Biochemistry (21 decks)

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