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Nucleic Acid Biochem I > Nucleic Acids Structure, supercoiling and degradation > Flashcards

Nucleic Acids Structure, supercoiling and degradation Flashcards

Readings: 305 - 316

1
Q

There are 2 bonds between what base pair?

A

Adenine and thymine

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2
Q

There are 3 bonds between what base pair?

A

Guanine and cytosine

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3
Q

How many Angstroms (Å) is the length of one helical turn in DNA?

A

34

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4
Q

How many base pairs are in one helical turn of DNA?

A

10.5

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5
Q

What is the major groove between in DNA?

A

Helical turns

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6
Q

What is the minor groove in DNA structure?

A

Between strands

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7
Q

What is the width of DNA?

A

20 Å

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8
Q

What is the distance between each base pair?

A

3.4 Å

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9
Q

What two things stabilizes DNA double helix?

A

Hydrogen bonds and base stacking

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10
Q

When is A-DNA formed?

A

In high salt and low humidity situations

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11
Q

What is the helix sense for B-DNA, A-DNA and Z-DNA?

A

B-DNA: Right handed
A-DNA: RIght handed
Z-DNA: Left handed

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12
Q

What is the sugar pucker conformation for B-DNA, A-DNA and Z-DNA?

A

B-DNA: C-2’ endo
A-DNA: C-3’ endo
Z-DNA: C-2’ endo for pyrimidines

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13
Q

What is the glycosyl bond conformation for B-DNA, A-DNA and Z-DNA?

A

B-DNA: Anti
A-DNA: Anti
Z-DNA: Anti for pyrimidines and syn for purines

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14
Q

What is the structure of Z-DNA?

A

Sequence of alternating purines and pyrimidines, with Z-DNA binding proteins attached

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15
Q

What are the 3 conformations of DNA in order of most compact to least compact? (smallest grooves to biggest grooves)

A

A form
B form
Z form

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16
Q

What do B, A and Z DNA’s look looking ‘into’ them?

A

B: Center disk with spokes inside a circle
A: A circle within a circle
Z: A star in a circle

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17
Q

Describe a nucleoside with a syn glycosidic bond

A

The base points in the same direction as the pentose, that is, it is right over it.

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18
Q

Describe a nucleoside with a anti glycosidic bond

A

The base points away from the pentose ring

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19
Q

Which conformation of glycosidic bond is favoured in DNA? Why?

A

The anti conformation is favoured because the syn conformation has more steric hindrance due to crowding of the pentose ring

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20
Q

What is linking number (Lk)?

A

The number of helical turns in DNA

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21
Q

How can linear DNA form supercoils?

A

linear DNA between two anchored points can supercoil

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22
Q

What is LKo?

A

LK in relaxed DNA

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23
Q

How can LKo be determined?

A

By dividing the number of base pairs in strand by 10.5 (remember 10.5 bp per helical turn of B-DNA)

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24
Q

What is S for nucleic acids?

A

Number of supercoils in DNA

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25
Q

How can number of supercoils (S) be determined for DNA?

A

S = LK - LKo

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26
Q

What is ethidium bromide used for?

A

Used for visualizing DNA when exposed to UV, but also highly mutagenic. It intercalates in DNA like a base pair.

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27
Q

How does ethidium bromide effect supercoiling?

A

It decreases negative supercoiling

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28
Q

What is the LKo equation for ethidium bromide’s effect on DNA?

A

LKo = (# bp) / (10.5 + # of ethidium bromide)

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29
Q

Tou have a strand of circular DNA 200 helical turns (Lk = 200) and it is relaxed (Lko = 200). What does Lk become if DNA is negatively supercoiled by two helical turns?

A

Lk = 198 with a ΔLk of -2

30
Q

How do positive supercoils affect the number of helical turns (Lk)?

A

They increase the number of helical turns

31
Q

How do negative supercoils affect the number of helical turns (Lk)?

A

They decrease the number of helical turns

32
Q

Does supercoiled DNA Lk decrease or increase further from a well in a western blot? Why?

A

Lk decreases further along a western blot, as DNA is more supercoiled further along a western blot.

33
Q

What does topoisomerase I do?

A

Relaxes negative supercoils

34
Q

What does topoisomerase II (DNA gyrase) do?

A

Introduces negative supercoils

35
Q

What does Topoisomerase III do? Which class is it?

A

Specialized functions in DNA repair and replication. Type 1 topoisomerase

36
Q

What does type I topoisomerase do?

A

Relaxes negative supercoils

37
Q

What does type II topoisomerase do?

A

Introduces negative supercoils

38
Q

What effect will type II topoisomerase have on the Lk of DNA?

A

Decrease Lk by introducing negative supercoils

39
Q

What effect will type I topoisomerase have on the Lk of DNA?

A

Increase Lk by relaxing negative supercoils (will not relax above Lko)

40
Q

How many helical turns does type I and topoisomerase I deal with in a single action? How many strands at once?

A

I turn per action along a single strand

41
Q

How many helical turns does type II topoisomerases make in a single action?

A

2 turns per action.

42
Q

What are nucleases?

A

Enzymes that catalyze the hydrolysis of phosphodiester bonds in nucleic acids

43
Q

Name 6 varieties of nuclease

A
  • ribonuclease (RNase)
  • Deoxyribonuclease (DNase)
  • Non specific nuclease
  • Single strand nuclease
  • Double strand nuclease
  • Single and double strand nuclease
44
Q

What are two modes of attack for nucleases?

A
  • Endonuclease activity, hydrolyzes interior phosphodiester bonds
  • Exonuclease activity, acts sequentially from one end to the other (ie. 5’ to 3’ or 3’ to 5’)
45
Q

Give an example of two nucleases used in food digestion

A

Pancreatic DNase I and RNase A

46
Q

how are nucleases used in gene regulation?

A

RNA turnover

47
Q

What are the products of RNase phosphodiester bond cleavage?

A

A 5’ OH and 3’ Phosphate

48
Q

What are the products of DNase phosphodiester bond cleavage?

A

A 3 OH and 5’ phosphate

49
Q

What are three types of base specificity in nucleases?

A
  • Specific for a single base (eg. RNase T1 specific of G)

- Specific for a particular class of base (eg. RNase S, specific for pyrimidines

50
Q

True or false, alkalis hydrolyze both RNA and DNA

A

False, alkalis hydrolyze only RNA

however, Alkalis denature DNA

51
Q

What configuration is double stranded RNA in?

A

A-DNA conformation

52
Q

What are pucker conformations? List two

A

Pentose conformations (it’s not flat).

  • C3’-endo, where 3’ carbon is protruding up
  • C2’-endo, where 2’ carbon is protruding up
53
Q

What is the norm for supercoils in nature?

A

Negative supercoils

54
Q

Can loci in the genome ever have positive supercoils?

A

Yes, though temporarily

55
Q

Why do thermophilic bacteria have positive supercoils?

A

Because it makes DNA more stable and able to withstand high temperatures

56
Q

Which topoisomerase has a ‘controlled relaxation?’

A

Type I Topoisomerases

57
Q

What is another name for topoisomerase II in prokaryotes?

A

DNA gyrase

58
Q

Which type of topoisomerase requires ATP?

A

Type II, because it introduces negative supercoils, goes away from relaxed (lower energy) state

59
Q

What type of topoisomerase can separate catenated chromosomes?

A

Type IV

60
Q

What is the conventional direction of the top strand of DNA (when drawn)?

A

5’ to 3’

61
Q

What phosphates are cyclic, what do they become susceptible to? What are results of this?

A

Susceptible to hydrolysis by water, making monophosphate intermediates (2’ and 3’)

62
Q

Why can’t alkali solutions hydrolyze DNA?

A

Because it lacks the 2’ OH group that RNA has

63
Q

What is topology with supercoiling?

A

A branch of mathematics that studies the properties of an object that do not change under continuous deformations.

64
Q

Is most cellular DNA underwound or overwound?

A

Underwound, where the molecule has fewer helical turns than expected for B-DNA

65
Q

If an 84 bp segment of circular DNA had one of its helical turns removed, how many base pairs would there be for every helical turn?

A

84 bp/10.5 bp per turn = 8 helical turns

84 bp/7 helical turns = 12 bp per turn

66
Q

Explain why supercoiling occurs, thermodynamically

A

When there are more or less helical turns in a molecule of DNA, the molecule is more unstable and thermodynamically strained. The strain is accommodated by coiling the axis on itself.

67
Q

How does topoisomerase change linking number at a molecular level?

A

It breaks one strand of DNA, rotates the end of it 360 degrees about the unbroken strand and rejoins the broken ends.

68
Q

What is writhe (Wr)?

A

A measure of the coiling of the helical axis

69
Q

What is twist (Tw)?

A

The local twisting or spatial relationship of neighbouring base pairs

70
Q

What is the equation associated with Lk, Wr and Tw? Why?

A

Lk = Tw + Wr

Because when linking number changes, some strain is compensated with twist and some is compensated with writhe

71
Q

What form of supercoiling is observed in isolated DNA?

A

plectonemic supercoiling, extended right hand coiling (like telephone knots)

72
Q

What are two forms of compaction from supercoiling? Which compacts more and gives chromosomes their density?

A
  • Plectonemic (most stable in solution)
  • Solenoidal (eg. around nucleotides)

Solenoidal supercoiling compacts more. These forms are readily interconvertible

Nucleic Acid Biochem I (5 decks)

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