4. DNA Synthesis

Question 1

Describe how the DNA helix facilitates DNA replication (4pts)

Answer 1

1. Two antiparallel polynucleotide form a right handed helix in which the bases are on the inside whilst the sugars and phosphates are on the outside.
2. B form DNA so there are 20 bases per helical turn
3. Polynucleotide chains linked together by hydrogen bonds between bases- A+T, C+G therefore one strand in complementary in sequence to the other.
4. DNA synthesis requires unwinding and opening followed by the copying of each strand.

Question 2

Describe Semi-conservative DNA replication (2 pts)

Answer 2

1. DNA replication is semi- conservative which means it consists of one parental DNA strand and one new strand.
3. Parent DNA opens up
4. Replication machinery opens up the parental DNA strands and copies the strands to synthesise a new complementary strand.

Question 3

Describe a replication origin (3pts)

Answer 3

1. DNA replication is initiated at specific sites on DNA called replication origin.
2. Replication origions are recognised by an initiation complex.
3. DNA at the origin unwinds to form a replication bubble and allows access to the replication machinery.

Question 4

Describe the bacterial cell cycle (4m)

Answer 4

1. Lasts for 20-30 mins
2. Very quick
3. Daughter cells go into the s phase of the cell cycle
4. Largely occupied by synthesis of the DNA.

Question 5

Describe the mammalian cell cycle (4pts)

Answer 5

• division lasts for 16-24 hours

1. Mitosis- chromosomes are condensed. Produced 2 daughter cells.
2. G1 phase- cells are preparing to replicate their DNA to synthesise their DNA in the s phase.
3. S phase- DNA synthesis occurs. 8 hours.
4. G2 phase- DNA and cells are being prepared for condensation into chromosomes.

Question 6

Describe bacterial DNA Replication (5pts)

Answer 6

1. Bacterial DNA chromosomes are condensed, circular and supercoiled.
2. The chromosomes have a single origin of replication. Proteins can bind at this origin which results in the unwinding of 2 parental strands at this location. This allows the initiation of the separation of the parental strands to begin and generates a replication bubble.
3. Under the replication bubble there are two replication forks on either side of the bubble.
4. Replication in bacteria is bidirectional- occurs in both directions.
5. Bacterial DNA has a single replication origin- replicates in 20-30 mins.

Question 7

Describe the steps in Bacterial DNA replication (3pts)

Answer 7

1. At the replication forks the helix unwinds and moves through the DNA on both sides of the replication forks.
2. Over time the replication forks gradually moves through the circular DNA and do this on both sides of the replication bubble..
3. Over a short period the parental strands would separate and be copied to make the daughter chromosomes.

Question 8

Describe DNA replication in eukaryotic cells (3pts)

Answer 8

• In eukaryotic cells, DNA replication initiates at multiple replication origins.
  1. Steps:

1. The parental DNA helix unwinds and is copied to make the newly synthesised DNA.
2. Over time the replication forks move away to either side of the replication bubble causing the replication bubble to get larger.
3. Eventually the newly synthesised DNA will all be joined up and we end up synthesising the 2 new daughter chromosomes.

Question 9

Describe DNA polymerase (5m)

Answer 9

1. Enzyme which makes polynucleotide chains.
2. Acts in the 5 prime to 3 prime direction only- DNA polymerase all add nucleotides to the 3 prime OH end of the newly synthesised strand which means they elongate in the 5 prime to 3 prime direction.
3. Utilises A-T and C-G base pairing to synthesise a new DNA strand.
4. Has a proof- reading function- goes back and checks nucleotides, If there is a wrong nucleotide in the newly synthesised strand it erases it and inserts the correct nucleotide.
5. Cells have multiple DNA polymerase-

bacteria= 5 different polymerases 
eukaryotes= 15 different DNA polymerases

Question 10

Describe the leading and lagging strand (3pts)

Answer 10

1. Leading strand= synthesis occurs so that you have a continuous copying of the parental DNA strand
2. Lagging strand= synthesised in a discontinuous way and forms small Okazaki fragments of DNA that have to be processed and then linked together to make the complete joined up strand.
3. DNA polymerase can only synthesise in the 5 prime to 3 prime direction. This means that while the leading strand is continually synthesised with no interruptions at all after the first primer is placed the lagging strand is synthesised at intervals as DNA polymerase can only synthesise in one direction and since both template strands are anti parallel one will be the wrong way round.

Question 11

Describe Telomerase ( 2 pts)

Answer 11

1. Reverse transcriptase enzyme expressed in stem and cancer cells
2. Synthesise DNA at the ends of chromosomes on telomeres to offset DNA loss during normal replication.

Question 12

Describe repair Polymerase (1pt)

Answer 12

Repair DNA damage

Question 13

Describe Translesion polymerase (1pt)

Answer 13

Allows replication forks to bypass damage that cannot be repaired

Question 14

Describe DNA synthesis in terms of the strands (3pts)

Answer 14

1. As the template strand unwinds and becomes more available the leading strand is able to be continually copied by the DNA polymerase and travels towards the same direction of the replication fork.
2. On the lagging strand the DNA is made discontinuously in Okazaki fragments.
3. Okazaki fragments have to be processed to give a newly synthesised strand.

Question 15

Describe DNA replication (2 pts)

Answer 15

• DNA replication is a multi enzyme process

1. DNA helices acts at the replication fork and uses ATP to break the base pairs between the two strands of the parental DNA
2. This opens up the DNA allowing it to be copied by the replicative polymerase.
   - To prevent DNA from being digested by nucleases it is coated with a DNA binding protein called single stranded binding protein.

Question 16

Describe DNA replication in bacteria ( 5 pts)

Answer 16

1. Pul 3 replicates the leading strand and lagging strand.
2. Primase is an enzyme on e.coli which provides a primer that is used by Pul 3 to copy the DNA strand on the lagging strand.
3. This produces Okazaki fragments where each fragment begins with RNA and the rest of it is DNA.
4. When processing the Okazaki fragments the RNA component had to be removed by repair DNA polymerase which replaces RNA with DNA.
5. This produces DNA fragments which can be ligated together by the enzyme Ligase.

Question 17

Describe DNA replication in eukaryotes (4pts)

Answer 17

1. Polymerase Alpha in conjunction with primase lays down a RNA-DNA primer on the lagging strand.
2. The DNA-RNA primer is then used by polymerase to make more DNA.
3. The polymerase delta as it synthesises DNA will encounter the previous Okazaki fragments and will displace the RNA.
4. This produces DNA fragments which can be ligated together by the enzyme DNA ligase.

Question 18

Why does DNA replication have high fidelity (3pts)

Answer 18

1. Base pairing in DNA, proof-reading by DNA polymerase and the mismatch repair system ensures that DNA replication proceeds with high fidelity.
2. DNA polymerase has a very low error rate due to base pairing and the proof reading function of the enzyme.
3. The mismatch repair system corrects most of the errors. The multi-enzyme system is highly conserved,

Question 19

Describe clinically important drugs (3pts)

Answer 19

1. Antibacterial drugs= Gyrase inhibitors
2. Anti-tumour drugs= topo II inhibitors
3. Antiviral AIDS= Reverse transcriptase inhibitors.