3(b)

Question 1

What happens in elongation part of DNA replication?

Answer 1

DNA polymerase adds DNA nucleotides to the 3’ end of the newly synthesized polynucleotide strand. The template strand specifies which of the four DNA nucleotides (A, T, C, or G) is added at each position along the new chain. Only the nucleotide complementary to the template nucleotide at that position is added to the new strand

Question 2

What does the replisome include? (8 things)

Answer 2

Sliding clamp,leading DNAP,Topoisomerase,Helicase,Primase,Lagging DNAP,Single strand binding proteins SSBP,Okazaki fragments.

Question 3

Which end can DNAP add nucleotides to?

Answer 3

DNA polymerase cannot initiate new strand synthesis; it only adds new nucleotides at the 3’ end of an existing strand.

Question 4

DNAP will not bind ssDNA so what is added?

Answer 4

erasable RNA primers, one on leading and many on the lagging strand.

Question 5

What does primase do?

Answer 5

Initiates the polypeptide synthesis by creating short RNA polypeptide strands complimentary to the template DNA strand. This short stretch of RNA is called the primer. Once it has been synthesised primase exits and DNAP extens the new strand with complimentary DNA nucleotides.Eventually, the RNA nucleotides in the primer are removed and replaced with DNA nucleotides.

Question 6

In bacteria what removes primers? Is this the part that is removed during hydrolysis o make the klenow fragment?

Answer 6

DNAP-1 has own primer moving nuclease activity.Yes this is the part that is removed by hydrolysis to make the Klenow fragment.

Question 7

Why is the klenow fragment that is part of the bacterial NAP-1 repair polymerase a model system? (3)

Answer 7

• it is present in high copy number
• it retains the 5’ → 3’ polymerase activity and the 3’ → 5’ exonuclease activity for removal of precoding nucleotides and proofreading, but loses its 5’ → 3’ exonuclease activity
• the Klenow fragment, which lacks this activity (exonuclease) can be very useful in research

Question 8

What can klenow fragment be used in? (4)

Answer 8

• synthesis of double-stranded DNA from single-stranded templates
• Filling in receded 3’ ends of DNA fragments to make 5’ overhang blunt
• Digesting away protruding 3’ overhangs
• Preparation of radioactive DNA probes

Question 9

is leading strand synthesized towards or away from replication fork?

Answer 9

continuously toward the replication fork as helicase unwinds the template double-stranded DNA.

(the two newly-synthesized strands grow in opposite directions because the template strands at each replication fork are antiparallel)

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Question 10

What are the fragments called that are synthesized in the direction away from replication fork?

Answer 10

Okazaki fragments, and each fragment begins with its own RNA primer.

Question 11

In eukaryotes when does DNAP halt and drop off?

Answer 11

in Termination when it it reaches a section of DNA template that has already been replicated.It cannot catalyze the formation of a phosphodiester bond between the two segments of the new DNA strand.

Question 12

What are nicks?

Answer 12

unattached sections of the sugar-phosphate backbone in an otherwise full-replicated DNA strand.

Question 13

Enzymes that remove RNA primers?

Answer 13

FEN1 (flap endonulcease 1) and RNase H

Question 14

Where do FEN1 and RNase remover primers from? and what is left?

Answer 14

FEN1 and RNase H remove RNA primers at the start of each leading strand and at the start of each Okazaki fragment, leaving gaps of unreplicated template DNA

Question 15

Once primers are removed what happens in eukaryotes?

Answer 15

1) a free-floating DNA polymerase lands at the 3’ end of the preceding DNA fragment and extends the DNA over the gap
2) the enyzme ligase joins the sugar-phosphate backbones at each nick site and after all nicks are connected the daughter DNA is complete.

Question 16

Where does termination occur in eukaryotes?

Answer 16

telomeres,DNAP cannot replicate telomeres at very 3’ end of eukaryotic linear chromosome.

Question 17

what is telomerase and what cells possess it?

Answer 17

Telomerase is a RT with an internal RNA template which extends telomeres in germ-line cells

Question 18

How many DNAP needed in eukaryotes per dna strand?

Answer 18

At replication fork there are two molecules of DNA polymerase and a large number of other essential proteins

Question 19

which direction are the lagging and leading strands synthesised in?

Answer 19

Lagging strand is synthesised discontinuously in discrete 5′→3′ chunks
whilst the Leading strand is synthesised continuously in 5′→3′ direction

Question 20

do both prokaryotic and eukaryotic cells contain several different DNA polymerases that play distinct roles in the replication and repair of DNA?

Answer 20

yes

Question 21

Do prokaryotes have 1 type of DNAP?

Answer 21

No

Question 22

what revealed the multiplicity of DNAP?

Answer 22

the isolation of a mutant strain of E. coli that was deficient in polymerase I

Question 23

What DNAP do eukaryotic cells have?

Answer 23

five DNA polymerases: α, β, γ, δ, and ε. Polymerase γ is located in mitochondria and is responsible for replication of mitochondrial DNA. The other four enzymes are located in the nucleus and are therefore candidates for involvement in nuclear DNA replication.

Question 24

What are DNA polymerases α, δ, ε and β used in?

Answer 24

Polymerases α, δ, and ε are most active in dividing cells, suggesting that they function in replication. In contrast, polymerase β is active in nondividing and dividing cells, suggesting that it may function primarily in the repair of DNA damage

Question 25

What does the primosome do and what type of organism is it found in?

Answer 25

a protein complex responsible for creating RNA primers on single stranded DNA during DNA replication.

The primosome consists of seven proteins: DnaG primase, DnaB helicase, DnaC helicase assistant, DnaT, PriA, Pri B, and PriC.

It is found in bacteria such as E.coli

Question 26

What does DNAP alpha do? and what is it also known as?

Answer 26

It is also known as Pol α and is an enzyme complex found in eukaryotes that is involved in initiation of DNA replication

Question 27

How does Pol alphas Pol α limited processivity and lack of 3′ exonuclease activity for proofreading errors reflect its role?

Answer 27

It is not well suited to accurately copying long templates (unlike Pol Delta and Epsilon). Instead Pol α is responsible for the initiation of DNA replication at origins of replication (on both the leading and lagging strands) and during synthesis of Okazaki fragments on the lagging strand.