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CMMB 411 > Topic 3 (DNA Structure and Topology) > Flashcards

Topic 3 (DNA Structure and Topology) Flashcards

1
Q

Where do purines attach to the ribose sugar?

A

C1, N9 of the base

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2
Q

Where do pyrimidines attach to the ribose sugar?

A

C1, N1 of the base

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3
Q

Which two interactions stabilize the structure of DNA?

A

Hydrogen bonds between the bases and pi-stacking of the bases

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4
Q

What effect does pi-stacking have that stabilizes the structure of DNA?

A

Hydrophobic effect and Van der Waals interactions

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5
Q

What is the handedness of a B-form DNA double helix?

A

Right-handed

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6
Q

What is the handedness of an A-form DNA double helix?

A

Right-handed

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7
Q

What is the handedness of a Z-form DNA double helix?

A

Left-handed

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8
Q

What form of the dNTPs is found in B-form DNA (anti or syn)?

A

Anti

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9
Q

What form of the dNTPs is found in Z-form DNA (anti or syn)?

A

Syn

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10
Q

Which direction does right-handed DNA spin?

A

Clockwise

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11
Q

What is the tautomer of an amide?

A

Imide

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12
Q

What is the tautomer of an enol?

A

Keto

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13
Q

What is tautomerization?

A

The migration of a hydrogen atom between areas on the same molecule

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14
Q

Define a tautomeric shift

A

The spontaneous rearrangement of nitrogenous bases that allow for hydrogen bonding of mismatch pairs

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15
Q

If a T tautomerized into its enol form, what would it pair with?

A

G

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16
Q

If a C tautomerized into its enol form, what would it pair with?

A

A

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17
Q

Base flipping allows for:

A

Homologous recombination and DNA repair

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18
Q

Explain the Mica experiment

A

A strand of DNA was attached to mica and exposed to DNase I. The enzyme would make a cut in the strand not attached to the mica every 10 nucleotides or so

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19
Q

What three strategies were used in the Mica experiment?

A

Affinity (attachment of DNA to mica), restriction digest, and gel electrophoresis

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20
Q

What conclusions could be made from the Mica experiment?

A

The DNA makes a 360 degree rotation every ~10.5 nucleotides, or every nucleotide is twisted 36 degrees from the previous one

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21
Q

What is the length of a B-DNA strand relative to the other forms?

A

Normal

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22
Q

What is the length of a A-DNA strand relative to the other forms?

A

Short

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23
Q

What is the length of a Z-DNA strand relative to the other forms?

A

Long

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24
Q

How many base pairs per helical turn for B-DNA?

A

10.5

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25
Q

How many base pairs per helical turn for A-DNA?

A

11

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26
Q

How many base pairs per helical turn for Z-DNA?

A

12

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27
Q

How many degrees per residue for B-DNA?

A

36

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28
Q

How many degrees per residue for A-DNA?

A

33

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29
Q

How many degrees per residue for Z-DNA?

A

30

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30
Q

True/False? Diffraction pattern lines are rotated 45 degrees to the actual lines formed by the DNA

A

False. They are perpendicular

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31
Q

What is pitch?

A

The length between rotations, 34 Angstroms

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32
Q

What is rise?

A

The distance between each nucleotide, 3.4 Angstroms

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33
Q

What is the radius of a DNA double helix?

A

10 Angstroms

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34
Q

When B-form DNA was used for x-ray diffraction, why did Franklin and Gosling conclude that DNA was a double helix?

A

Spot 4 was missing due to destructive interference because the phosphate backbones overlapped with each other

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35
Q

Why is the “X” formed in photograph 51 not formed by perpendicular lines?

A

Major and minor grooves

36
Q

Define denaturation of DNA

A

The disruption of hydrogen bonds by heat, pH, or [salt]

37
Q

Where can Tm (melting temperature) be found on a Temperature vs. Absorbance graph?

A

The midway point between where the absorbance changes from what you’d expect to see in double-stranded DNA to what you’d expect to see in single-stranded DNA (half of the DNA is denatured)

38
Q

What does Tm represent?

A

The temperature at which the sample consists of 50% dsDNA and 50% ssDNA

39
Q

Which factors affect Tm?

A

G:C content

Ionic strength of the solution

40
Q

What effect does increasing G:C content have on TM? Why?

A

Increase; 3 hydrogen bonds require more energy than 2 (A:T), and G:C have lower entropy

41
Q

What effect does increasing salt content have on TM? Why?

A

Increases in [salt] stabilize the phosphate backbone by decreasing the repulsive force between the two backbones, so dsDNA is more stable than usual

42
Q

What absorbance does DNA absorb UV?

A

260nm

43
Q

Describe the hyperchromic effect

A

ssDNA absorbs >40% of UV at 260nm than dsDNA due to dsDNA’s base stacking

44
Q

What effect does base stacking have on UV absorbance?

A

Decreases absorbance

45
Q

How does hybrid dsDNA occur?

A

When re-annealing, ssDNA can associate with another strand that has a similar sequence to form hybrid DNA

46
Q

When 2 different but similar dsDNAs are denatured and re-annealed, what are the possible outcomes?

A

Could result in 2 hybrid strands or 2 normal strands

47
Q

What experimental methods is DNA hybridization important for?

A

Southern blots, Northern blots, DNA and RNA microarrays, next-gen sequencing, identification of mutations

48
Q

What is cccDNA?

A

Covalently closed, circular DNA

49
Q

What is an example of cccDNA?

A

Bacterial plasmids

50
Q

What can linear DNA do that circular DNA can’t?

A

Can freely rotate and unwind

51
Q

During transcription, which end of the DNA is overwound? Why?

A

5’ end of the strand that is being transcribed. This is because as the DNA unwinds so RNA pol can transcribe, extra torsional stress is put on the end that it moves towards

52
Q

During transcription, which end of the DNA is underwound? Why?

A

3’ end of the strand that is being transcribed. This is because as the DNA unwinds so RNA pol can transcribe, torsional stress is released from the end it moves away from

53
Q

What is the formula for linking number (Lk)

A

Twist (Tw) + Writhe (Wr)

54
Q

What does one twist and one writhe count as in terms of linking number?

A

Both -1

55
Q

Define linking number

A

An integer representing the number of times it takes for a strand of DNA to pass through the other strand in order for the two strands to be entirely separated

56
Q

True/False? Topoisomers differ in both linking number and base order

A

False. Only linking number

57
Q

Using gel electrophoresis, how can we tell which band represents the relaxed coil or the supercoil? Why?

A

The smaller band is the supercoil as it is more compact and thus has less frictional resistance

58
Q

List the order in which you would find a linear piece of DNA, supercoil with a Lk of -5, relaxed coil, and a supercoil with a Lk of -2 resulting from gel electrophoresis from largest to smallest

A

Relaxed, linear, supercoiled (-2), supercoiled (-5)

59
Q

True/False? Supercoiling is determined by writhe, not twist

A

True

60
Q

Which type of supercoil (positive or negative) is human DNA usually found in? Why?

A

Negative; It requires less energy to unwrap which is necessary for regular gene transcription

61
Q

Which type of supercoil (positive or negative) is extremophile DNA usually found in? Why?

A

Positive; It requires more energy to unwrap (more stable), so DNA is less likely to be damaged or denatured by extreme environment

62
Q

Define supercoiled DNA

A

DNA that twists upon itself because it is overwound or underwound (and thereby strained) relative to B-DNA

63
Q

True/False? Topoisomerases can both introduce and relieve supercoils

A

True

64
Q

Which direction must DNA be wound in to result in a negative supercoil?

A

Counterclockwise

64
Q

Which direction must DNA be wound in to result in a positive supercoil?

A

Clockwise

65
Q

A DNA strand has an Lk of -3 and is negatively coiled. What must be done to return the DNA to its relaxed form (Lk=0)?

A

A topoisomerase must break both strands of DNA, the DNA must then be rotated 360 degrees 3 times clockwise

66
Q

A negatively supercoiled DNA (Lk=-3) must be unwound to its relaxed form (Lk=0). Explain the steps

A

Topoisomerase must break the DNA so one helix can rotate 3x360 degrees clockwise ***

67
Q

How many times does a supercoiled DNA cross over itself if its Lk=-4? How many loops does it have?

A

Crosses over 4 times, has 5 loops

68
Q

Which direction does a negative supercoil loop around a histone core?

A

Clockwise

69
Q

Which direction does a positive supercoil loop around a histone core?

A

Counterclockwise

70
Q

What are the functions of supercoiling?

A

To reduce the space and allow for DNA to be packaged into a small nucleus

To prevent or resolve DNA entanglement

Positive supercoiling protects DNA from thermal denaturation and regulates gene expression in extreme conditions

71
Q

True/False? Negative supercoiling stores free energy required to facilitate strand compaction

A

False. Facilitates strand separation as negative coiling loosens up DNA winding

72
Q

True/False? Topoisomerases do double-stranded breaks

A

False. Topoisomerase II does double, topoisomerase I does single stranded breaks

73
Q

What is a type I topoisomerase?

A

It makes a ssDNA cut, does not require energy, and relaxes/introduces supercoils by 1 per digestion

74
Q

Which amino acid residue on Topo I attacks the DNA?

A

Tyrosine (Tyr, Y)

75
Q

Describe the steps performed by Topo I

A

DNA is nicked by attack of tyrosine

The other side of the cleaved DNA is held by the enzyme

The uncleaved strand passes through the opening

DNA ligation (no ATP required)

DNA is released

76
Q

Why is energy not needed for Topo I to function?

A

Topo I is covalently linked to DNA after the nicking step. At the start of the reaction, one phosphate ester bond is broken and another one is formed while the strands rejoin, so the number of phosphate bonds remains the same

77
Q

What is a type II topoisomerase?

A

Makes dsDNA cuts, requires energy, and reduces/introduces supercoils 2 per digestion

78
Q

What is a common name for Topo II?

A

Gyrase

79
Q

True/False? Gyrase can be found in both prokaryotes and eukaryotes

A

False. Only prokaryotes

80
Q

What allows Topo I to do a catenation/decatenation?

A

ssDNA

81
Q

What reactions is Topo II useful for?

A

Catenation/decatenation of dsDNA, and separating entangled DNA after replication

82
Q

As the length of a reaction between topoisomerase and DNA goes on, what happens? What do we see in gel electrophoresis?

A

Lk decreases, “supercoiled” band moves closer and closer to the “relaxed” band

83
Q

What is ethidium bromide (EtBr)?

A

A carcinogenic dye used to stain DNA which intercalates between stacked nucleotide bases, decreasing the angle per base from 36 degrees to 10 degrees

84
Q

Why is EtBr carcinogenic?

A

Because it opens up the DNA, it is more susceptible to radiation/mutations

85
Q

True/False? EtBr increases the length of DNA

A

True

CMMB 411 (8 decks)

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