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

Lecture 22. DNA II Flashcards

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
What is the first rule and conclusion concerning DNA polymerase
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
26
What is a primer strand ?
A pre-existing strand
27
What can DNA polymerase only add nucleotides to ?
A primer strand which is already base-paired with a template strand
28
What is the second rule and conclusion concerning DNA polymerase ?
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
29
What direction does DNA synthesis occur ?
In a 5' to 3' direction
30
What is a replication fork ?
The junction of an unwound DNA strand
31
What is the process of DNA replication ?
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
Is DNA replication continuous, semi- discontinuous or discontinuous ?
Semi discontinuous
33
How does the leading strand copy ?
Continuous, ie. a single long molecule
34
How does the lagging strand copy ?
In segments
35
What are the segments that the lagging strand copies in called ?
Okazaki fragments
36
What must eventually happen to Okazaki fragments ?
They must be joined up
37
How does the lagging strand copy ?
Discontinuously
38
What does the lagging strand require for discontinuous replication ?
Multiple primers
39
What are the three enzymatic activities performed by DNA polymerase I ?
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
What is the function of 3' to 5' exonuclease activity ?
Serves an error correcting or proof reading function. Removing any incorrectly matched bases, so that the polymerase can try again
41
What does DNAP I do if the incorrect base is added ?
DNAP I stalls
42
What is DNAP I very sensitive to ?
UV radiation damage
43
How many different DNAP's have been discovered in E. coli ?
5
44
What is the function of DNAP I ?
Replication and repair
45
What is the function of DNAP II ?
Repair
46
What is the function of DNAP III ?
The principal DNA replication enzyme
47
What is the function of DNAP IV ?
Repair
48
What is the function of DNAP V ?
Repair
49
What are some characteristics of DNA polymerase III ?
1. Very fast 2. Highly processive 3. Accurate 4. Small error rate
50
How fast is DNAP III ?
1000 bases added per second per enzyme
51
How many bases does DNAP III add before falling off the template ?
More than 500,000
52
How many errors does DNAP III make ?
1 error in 10 ^7 bases added
53
What is the final error rate of DNAP III ?
1 in 10 ^10
54
What does DNA polymerase always require to start the synthesis of a new strand ?
A base paired primer
55
What must the base paired primer that starts DNA replication be made of ?
RNA
56
What is the inhibitor of RNA polymerase ?
Rifampicin
57
What do okazaki fragments get started by ?
A short RNA primer
58
What are RNA primers made from ?
RNA primase
59
What is the RNA primer that primes the synthesis of the leading strand made from ?
The E. coli RNA polymerase RNA polymerase II
60
What are the only molecules that can incorporate ^3H-Uracil
RNA molecules
61
What do alkali treatments degrade ?
RNA
62
What evidence is there that the initiating primer is made of RNA ?
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
What is the function of RNA primase ?
Catalyses the synthesis of RNA primers
64
How are okazaki fragments joined together ?
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
What is nick translation ?
When DNAP I removes the RNA primer and replaces it with new DNA
66
What is the function of DNA ligase in the joining of okazaki fragments ?
Seals the nicks between DNA pieces
67
What do origins with adenine, thymine rich elements attract ?
Helicase and other replication enzymes
68
What is a telomere ?
The end of a chromosome
69
What is the function of a telomere ?
Protect chromosomes from damage
70
What is the composition of a telomere ?
Repetitive sequences of non-coding DNA
71
What is an indication of aging ?
Telomere shortening
72
What is the hayflick limit ?
Cells can only divide so many times
73
What are telomeres elongated by ?
Telomerase

Molecules to Cells - Genetics and Virology (16 decks)

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