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Molecules to Cells - Genetics and Virology > Lecture 22. DNA II > Flashcards

Lecture 22. DNA II Flashcards

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1
Q

What does replication via templating explain ?

A

Spontaneous variation - a key premise for Darwinian evolution

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2
Q

What are the three models (hypothesis) of DNA replication ?

A
  1. Semi-conservative replication
  2. Conservative replication
  3. Dispersive replication
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3
Q

What is semi conservative replication ?

A

Each strand acts as a template on which the other strand is synthesised

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4
Q

What is conservative replication ?

A

One helix contains the parent DNA strand while the other helix contains entirely new DNA (initiates synthesis of a new molecule while remaining intact)

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5
Q

What is dispersive replication ?

A

DNAs have alternating segments of both parental and newly-synthesized DNA interspersed on both strands

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6
Q

What is the Meselson-Stahl experiment ?

A

A centrifuge was used to separate DNA molecules labeled with isotopes of different densities

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7
Q

How does DNA replicate ?

A

Via template directed base pairing of nucleotides and step wise creation of phosphodiester bonds ?

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8
Q

How is DNA actually replicated (which hypothesis) ?

A

Semi conservative

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9
Q

What was the concept of replication forks first revealed by ?

A

Electron microscopy

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10
Q

What is an autoradiograph ?

A

An image on an x-ray film or nuclear emulsion produced by the pattern of decay emissions from a distribution of a radioactive substance

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11
Q

What is the requirement of DNA replication ?

A
  1. DNA polymerase

2. Nucleotide triphosphate

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12
Q

What are the three functions of enzymes in DNA replication ?

A
  1. Enzymes must allow DNA double helix to unwind
  2. Enzymes allow template driven copying
  3. Enzymes mediate error correction
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13
Q

What is the main replicative enzyme ?

A

DNA polymerase III

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14
Q

What are required for DNA polymerisation initiation ?

A

RNA primers

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15
Q

What is the function of helicase; a key enzyme in DNA replication ?

A

Unwinding/opening up DNA

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16
Q

What is the function of topoisomerases; a key enzyme in DNA replication ?

A

Dealing with over winding (positive supercoiling)

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17
Q

What is the function of polymerase and primase; a key enzyme in DNA replication ?

A

DNA replication

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18
Q

What is the function of polymerases; a key enzyme in DNA replication ?

A

Extend primers

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19
Q

Who discovered DNA polymerase I ?

A

Arthur Kornberg

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20
Q

What is the structure of DNA polymerase I ?

A

A single subunit protein

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21
Q

What did Kornberg devise to detect a DNA synthesis ?

A

An in vitro assay

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22
Q

What was in Kornberg’s assay mixture ?

A
  1. Total protein and cell extract from E. coli cells
  2. Template DNA
  3. Substrates required for DNA synthesis
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23
Q

What happened to Kornberg’s nucleotide triphosphates ?

A

They were radioactively labelled

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24
Q

Can DNA polymerase start a new DNA strand on its own ?

A

No

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25
Q

What is the first rule and conclusion concerning DNA polymerase

A

Rule: DNA polymerase can only add nucleotides to a pre-existing strand (primer) that is already base-paired with a template strand

Conclusion: DNA occurs via primer extension

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26
Q

What is a primer strand ?

A

A pre-existing strand

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27
Q

What can DNA polymerase only add nucleotides to ?

A

A primer strand which is already base-paired with a template strand

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28
Q

What is the second rule and conclusion concerning DNA polymerase ?

A

Rule: DNA can only add nucleotides to the 3’ end of a replicating strand

Conclusion: DNA synthesis is said to occur exclusively in a 5’ to 3’ direction

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29
Q

What direction does DNA synthesis occur ?

A

In a 5’ to 3’ direction

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30
Q

What is a replication fork ?

A

The junction of an unwound DNA strand

31
Q

What is the process of DNA replication ?

A
  1. Double stranded DNA must first unwind
  2. A new strand is formed by pairing complementary bases with an old strand
  3. Each replicated DNA molecule has one old and one new DNA strand
32
Q

Is DNA replication continuous, semi- discontinuous or discontinuous ?

A

Semi discontinuous

33
Q

How does the leading strand copy ?

A

Continuous, ie. a single long molecule

34
Q

How does the lagging strand copy ?

A

In segments

35
Q

What are the segments that the lagging strand copies in called ?

A

Okazaki fragments

36
Q

What must eventually happen to Okazaki fragments ?

A

They must be joined up

37
Q

How does the lagging strand copy ?

A

Discontinuously

38
Q

What does the lagging strand require for discontinuous replication ?

A

Multiple primers

39
Q

What are the three enzymatic activities performed by DNA polymerase I ?

A
  1. 5’ to 3’ DNA polymerizing activity (forward synthesis)
  2. 3’ to 5’ exonuclease activity (backwards, base error correcting)
  3. 5’ to 3’ exonuclease activity (forward, removal of RNA primers)
40
Q

What is the function of 3’ to 5’ exonuclease activity ?

A

Serves an error correcting or proof reading function. Removing any incorrectly matched bases, so that the polymerase can try again

41
Q

What does DNAP I do if the incorrect base is added ?

A

DNAP I stalls

42
Q

What is DNAP I very sensitive to ?

A

UV radiation damage

43
Q

How many different DNAP’s have been discovered in E. coli ?

A

5

44
Q

What is the function of DNAP I ?

A

Replication and repair

45
Q

What is the function of DNAP II ?

A

Repair

46
Q

What is the function of DNAP III ?

A

The principal DNA replication enzyme

47
Q

What is the function of DNAP IV ?

A

Repair

48
Q

What is the function of DNAP V ?

A

Repair

49
Q

What are some characteristics of DNA polymerase III ?

A
  1. Very fast
  2. Highly processive
  3. Accurate
  4. Small error rate
50
Q

How fast is DNAP III ?

A

1000 bases added per second per enzyme

51
Q

How many bases does DNAP III add before falling off the template ?

A

More than 500,000

52
Q

How many errors does DNAP III make ?

A

1 error in 10 ^7 bases added

53
Q

What is the final error rate of DNAP III ?

A

1 in 10 ^10

54
Q

What does DNA polymerase always require to start the synthesis of a new strand ?

A

A base paired primer

55
Q

What must the base paired primer that starts DNA replication be made of ?

A

RNA

56
Q

What is the inhibitor of RNA polymerase ?

A

Rifampicin

57
Q

What do okazaki fragments get started by ?

A

A short RNA primer

58
Q

What are RNA primers made from ?

A

RNA primase

59
Q

What is the RNA primer that primes the synthesis of the leading strand made from ?

A

The E. coli RNA polymerase RNA polymerase II

60
Q

What are the only molecules that can incorporate ^3H-Uracil

A

RNA molecules

61
Q

What do alkali treatments degrade ?

A

RNA

62
Q

What evidence is there that the initiating primer is made of RNA ?

A
  1. Okazaki fragments can incorporate ^3H-Uracil. Only RNA molecules that can incorporate ^3H-Uracil
  2. If okazaki fragments are treated with alkali, they get shorter. Alkali treatment only degrades RNA
63
Q

What is the function of RNA primase ?

A

Catalyses the synthesis of RNA primers

64
Q

How are okazaki fragments joined together ?

A
  1. DNAP III approaches the next RNA primer while making new okazaki DNA
  2. DNAP I takes over, removing the RNA primer and replacing it with new DNA (Called nick translation)
  3. DNA ligase seals the nicks between the pieces of DNA
65
Q

What is nick translation ?

A

When DNAP I removes the RNA primer and replaces it with new DNA

66
Q

What is the function of DNA ligase in the joining of okazaki fragments ?

A

Seals the nicks between DNA pieces

67
Q

What do origins with adenine, thymine rich elements attract ?

A

Helicase and other replication enzymes

68
Q

What is a telomere ?

A

The end of a chromosome

69
Q

What is the function of a telomere ?

A

Protect chromosomes from damage

70
Q

What is the composition of a telomere ?

A

Repetitive sequences of non-coding DNA

71
Q

What is an indication of aging ?

A

Telomere shortening

72
Q

What is the hayflick limit ?

A

Cells can only divide so many times

73
Q

What are telomeres elongated by ?

A

Telomerase

Molecules to Cells - Genetics and Virology (16 decks)

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