Organisation of DNA in chromatin - Professor Latchman

Question 1

What three ways can the differences in complexity between eukaryotic and prokaryotic DNA be shown?

Answer 1

1. The amount of DNA, E. coli has 4.2x10^6 and humans have 3x10^9. 4000 and 20000 genes respectively
2. The packing of DNA, Human DNA is packed around histone proteins in chromatin structure
3. The expression of genes is very different, compartmentalisation in eukaryotic cells means that paired transcription and translation cannot occur.

Question 2

What is the packing problem?

Answer 2

An individual cells DNA would stretch 2-metre longs in the double helix structure. This 2 metres of DNA needs to be condensed into the size of a chromosome which is about half a micron long

Question 3

What are the 5 different types of Histones and what is the Molar ratio between them?

Answer 3

H1                    1:
H2A                2:
H2B                2:
H3                   2:
H4                   2

Question 4

Describe the properties of Histone proteins

Answer 4

Histone proteins are rich in positively charged, basic amino acids such as Lysine and Arginine. The Histone proteins being basic is very important because the DNA is acidic due to the negative charges on the phosphate groups. Neutralising this negative charge with basic Histones allows for a more compact structure

Question 5

What is a Histone Octamer?

Answer 5

A Histone octamer is the structure of the core histone proteins that DNA wraps around twice. It consists of;
2 H2A
2 H2B
2 H3
2 H4
These 4 types of Histones are known as the core Histones

Question 6

What is a nucleosome?

Answer 6

The DNA is folded twice around a Histone Octamer consisting of two molecules of each of the 4 core Histones.

Question 7

What is chromatin?

Answer 7

DNA with the associated Histone proteins is called chromatin

Question 8

How did researchers work out the string of nucleosomes structure?

Answer 8

Researchers isolated chromatin and digested it with Micrococcal nucleases. Only a partial digestion was carried out as a full one would of digested the nucleosomes. Some Histone proteins are more susceptible to nucleases then others and this infers the location of these histones depending on where successful digestion had occurred. The histones that were more accessible to the nucleases appeared as peaks in a graph measuring absorbance. The researchers were amazed how they got regular peaks despite no sequence specificity. The micrococcal nucleases are much more likely to cut between nucleosomes rather then in the nucleosomes, a dot on a electron micrograph and the absorbance peaks show that a nucleosome has been cut on each side and that one nucleosome is about 200 base pairs in size.

Question 9

What happens if you leave the micrococcal nucleases for longer to digest?

Answer 9

The DNA that is within the nucleosome begins to be cut. If this occurs, you will end up with a core of 146 base pairs and this shows that there are 146 base pairs actually wrapped around the core complex and cannot be digested. The other 54 base pairs are in the linker region between 2 nucleosomes. When digestion occurs to this level the H1 protein is released.

Question 10

Why is the H1 Histone protein special?

Answer 10

It is the only type of Histone protein that is not in the Core Histones and it is not part of the core octamer and so it is released on sufficient digestion with nucleases. H1 is very important in the beads on a string stage of compactation. With H1 coordinated into the structure the DNA is much more compact.

Question 11

Describe how H1 allows the DNA to compact from 10nm thick to 30nm thick

Answer 11

H1 Histone protein has a globular structure with wings extending out from either side, these wings can interact with one another and pull the beads on a string structure together.

Question 12

Describe the two models that can explain how H1 allows the DNA to compact from 10 to 30nm thick

Answer 12

1. The solenoid or one-start-helix model, there are two nucleosome loops in each layer connected in the middle by linker region.
2. Zig-zag arrangement, The beads on a string structure are compacted and converted into a helical ribbon

Question 13

What is the last stage of compactation? after the helical structures have formed?

Answer 13

The helical DNA is thrown into a series of loops that range in size, this is called the euchromatin structure. The DNA goes from 30nm to 300nm thick. The areas of DNA that are not active are wrapped up much more tightly then areas that are active.
DNA does pack up more tightly then this during division when it goes into chromosomes, to make sure that division occurs correctly and that all the information goes into the daughter cells