DNA Replication

Question 1

Which end of the strand can be replicated?

Answer 1

Only adding to the 3’ OH end

Question 2

What is needed for Polymerisation of the nucleic acid chain?

Answer 2

1. dNTP’s
2. A template (parent strand to copy)
3. A primer (another strand paired with the template to give a 3’ end)

Question 3

How is semi-conservative replication described as?

Answer 3

semi-discontinuous

Question 4

How does DNA replication start?

Answer 4

Primed by RNA

• RNA primer is synthesised by primase
• DNA Polymerase III extends primer
• Process repeated to form ‘Okazaki’ fragments

(okazaki fragment about 1000-2000bp in length)

Question 5

How are primers removed during DNA replication?

Answer 5

• DNA polymerase I and Rnase H remove RNA and Pol I extends the DNA chain
• NIcks removed by DNA ligase

Question 6

How are the nicks closed between Okazaki fragments?

Answer 6

DNA ligase

Question 7

Which direction does DNA replication occur?

Answer 7

DNA replication is bidirectional so the replication fork is extended both ways

Question 8

what are the names of the different strands during replication?

Answer 8

• One new strand is made continuously known as the leading strand
• One strand is made in pieces which are then joined known as the lagging strand

Question 9

Where are the problems that occur when replicating at the ends of linear eukaryotic chromosomes?

Answer 9

• No problems at 3’ end as DNA Pol continues making strand until it falls off
• Problem at new 5’ end because the last RNA primer on the lagging strand end is removed but can’t be replaced

Question 10

What is telomerase?

Answer 10

An enzyme that adds a new sequence to compensate for the loss at 5’ end of lagging strand

-Telomerase is active in germ cells but not in most somatic cells

Question 11

What are the bacteria enzymes involved in DNA synthesis?

Answer 11

E. coli has 5 DNA polymerases:

1. DNA Pol I: removes RNA primer and replaces with DNA, also helps repair damaged DNA
2. DNA Pol II: restarting replication when blocked by damaged DNA, as a role in DNA repair
3. DNA Pol III: Chromosome replication
4. DNA Pol IV and DNA Pol V: allow replication to bypass some typed of DNA damage, also involved in DNA repair

Question 12

What does 3’-5’exonuclease do?

Answer 12

Involved in proofreading to check the nucleotide that has been inserted is correct

Question 13

What does Helicase do in DNA replication?

Answer 13

unwinds the DNA duplex

Question 14

What does the single-stranded DNA binding protein do?

Answer 14

Keeps the strands apart and helps prevent stem-loop formation

Question 15

What does primosome do?

Answer 15

Moves along with the lagging strand and contains DnaC and DnaG

Question 16

How does DNA Polymerase III work?

Answer 16

• DnaB unwinds the duplex
• Alpha subunits synthesise DNA
• Tau subunits ensures dimerisation of polymerase
• Beta clamp encircles DNA

Once the template is unwound, primase synthesises a RNA primer for lagging strand synthesis. The template is then pulled through the catalytic core allowing addition of nucleotides to the 3’ end

Question 17

What are the mammalian DNA Polymerases?

Answer 17

alpha - initial synthesis at RNA primer
delta - DNA replication of lagging strand
epsilon - DNA replication of leading strand
beta - repair
gamma - DNA synthesis at mitochondrion

Question 18

What are RNA primers removed by in mammals?

Answer 18

FEN1

Question 19

What restores the balance in supercoiled regions during DNA replication?

Answer 19

• Topoisomerase (can remove positive and negative supercoils but main function is to remove negative supercoils by tightening underwound molecules)
• DNA Gyrase (removes positive supercoils by relaxing overwound molecule)

Question 20

How long does it take for E. Coli to replicate?

Answer 20

DNA Pol III synthesises DNA at 900 nucleotides per second
-42 minutes per chromosome

But they can actually replicated between 20 and 35minutes because E. Coli starts to replicate again before the previous replication is complete