Nucleic Acids Structure, supercoiling and degradation

Question 1

There are 2 bonds between what base pair?

Answer 1

Adenine and thymine

Question 2

There are 3 bonds between what base pair?

Answer 2

Guanine and cytosine

Question 3

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

Answer 3

34

Question 4

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

Answer 4

10.5

Question 5

What is the major groove between in DNA?

Answer 5

Helical turns

Question 6

What is the minor groove in DNA structure?

Answer 6

Between strands

Question 7

What is the width of DNA?

Answer 7

20 Å

Question 8

What is the distance between each base pair?

Answer 8

3.4 Å

Question 9

What two things stabilizes DNA double helix?

Answer 9

Hydrogen bonds and base stacking

Question 10

When is A-DNA formed?

Answer 10

In high salt and low humidity situations

Question 11

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

Answer 11

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

Question 12

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

Answer 12

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

Question 13

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

Answer 13

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

Question 14

What is the structure of Z-DNA?

Answer 14

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

Question 15

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

Answer 15

A form
B form
Z form

Question 16

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

Answer 16

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

Question 17

Describe a nucleoside with a syn glycosidic bond

Answer 17

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

Question 18

Describe a nucleoside with a anti glycosidic bond

Answer 18

The base points away from the pentose ring

Question 19

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

Answer 19

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

Question 20

What is linking number (Lk)?

Answer 20

The number of helical turns in DNA

Question 21

How can linear DNA form supercoils?

Answer 21

linear DNA between two anchored points can supercoil

Question 22

What is LKo?

Answer 22

LK in relaxed DNA

Question 23

How can LKo be determined?

Answer 23

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

Question 24

What is S for nucleic acids?

Answer 24

Number of supercoils in DNA

Question 25

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

Answer 25

S = LK - LKo

Question 26

What is ethidium bromide used for?

Answer 26

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

Question 27

How does ethidium bromide effect supercoiling?

Answer 27

It decreases negative supercoiling

Question 28

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

Answer 28

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

Question 29

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?

Answer 29

Lk = 198 with a ΔLk of -2

Question 30

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

Answer 30

They increase the number of helical turns

Question 31

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

Answer 31

They decrease the number of helical turns

Question 32

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

Answer 32

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

Question 33

What does topoisomerase I do?

Answer 33

Relaxes negative supercoils

Question 34

What does topoisomerase II (DNA gyrase) do?

Answer 34

Introduces negative supercoils

Question 35

What does Topoisomerase III do? Which class is it?

Answer 35

Specialized functions in DNA repair and replication. Type 1 topoisomerase

Question 36

What does type I topoisomerase do?

Answer 36

Relaxes negative supercoils

Question 37

What does type II topoisomerase do?

Answer 37

Introduces negative supercoils

Question 38

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

Answer 38

Decrease Lk by introducing negative supercoils

Question 39

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

Answer 39

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

Question 40

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

Answer 40

I turn per action along a single strand

Question 41

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

Answer 41

2 turns per action.

Question 42

What are nucleases?

Answer 42

Enzymes that catalyze the hydrolysis of phosphodiester bonds in nucleic acids

Question 43

Name 6 varieties of nuclease

Answer 43

- ribonuclease (RNase) - Deoxyribonuclease (DNase) - Non specific nuclease - Single strand nuclease - Double strand nuclease - Single and double strand nuclease

Question 44

What are two modes of attack for nucleases?

Answer 44

- Endonuclease activity, hydrolyzes interior phosphodiester bonds - Exonuclease activity, acts sequentially from one end to the other (ie. 5' to 3' or 3' to 5')

Question 45

Give an example of two nucleases used in food digestion

Answer 45

Pancreatic DNase I and RNase A

Question 46

how are nucleases used in gene regulation?

Answer 46

RNA turnover

Question 47

What are the products of RNase phosphodiester bond cleavage?

Answer 47

A 5' OH and 3' Phosphate

Question 48

What are the products of DNase phosphodiester bond cleavage?

Answer 48

A 3 OH and 5' phosphate

Question 49

What are three types of base specificity in nucleases?

Answer 49

- Specific for a single base (eg. RNase T1 specific of G) | - Specific for a particular class of base (eg. RNase S, specific for pyrimidines

Question 50

True or false, alkalis hydrolyze both RNA and DNA

Answer 50

False, alkalis hydrolyze only RNA | however, Alkalis denature DNA

Question 51

What configuration is double stranded RNA in?

Answer 51

A-DNA conformation

Question 52

What are pucker conformations? List two

Answer 52

Pentose conformations (it's not flat). - C3'-endo, where 3' carbon is protruding up - C2'-endo, where 2' carbon is protruding up

Question 53

What is the norm for supercoils in nature?

Answer 53

Negative supercoils

Question 54

Can loci in the genome ever have positive supercoils?

Answer 54

Yes, though temporarily

Question 55

Why do thermophilic bacteria have positive supercoils?

Answer 55

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

Question 56

Which topoisomerase has a 'controlled relaxation?'

Answer 56

Type I Topoisomerases

Question 57

What is another name for topoisomerase II in prokaryotes?

Answer 57

DNA gyrase

Question 58

Which type of topoisomerase requires ATP?

Answer 58

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

Question 59

What type of topoisomerase can separate catenated chromosomes?

Answer 59

Type IV

Question 60

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

Answer 60

5' to 3'

Question 61

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

Answer 61

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

Question 62

Why can't alkali solutions hydrolyze DNA?

Answer 62

Because it lacks the 2' OH group that RNA has

Question 63

What is topology with supercoiling?

Answer 63

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

Question 64

Is most cellular DNA underwound or overwound?

Answer 64

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

Question 65

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?

Answer 65

84 bp/10.5 bp per turn = 8 helical turns 84 bp/7 helical turns = 12 bp per turn

Question 66

Explain why supercoiling occurs, thermodynamically

Answer 66

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.

Question 67

How does topoisomerase change linking number at a molecular level?

Answer 67

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

Question 68

What is writhe (Wr)?

Answer 68

A measure of the coiling of the helical axis

Question 69

What is twist (Tw)?

Answer 69

The local twisting or spatial relationship of neighbouring base pairs

Question 70

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

Answer 70

Lk = Tw + Wr Because when linking number changes, some strain is compensated with twist and some is compensated with writhe

Question 71

What form of supercoiling is observed in isolated DNA?

Answer 71

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

Question 72

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

Answer 72

- Plectonemic (most stable in solution) - Solenoidal (eg. around nucleotides) Solenoidal supercoiling compacts more. These forms are readily interconvertible