DNA replication

Question 1

the central dogma of biology

Answer 1

DNA to RNA = transcription
RNA to protein = translation

Question 2

importance of DNA replication

Answer 2

• ensures an exact copy of the species’ genetic info is passed from cell to cell
• if DNA failed to replicate itself meiosis and mitosis would pause

Question 3

features of DNA structure

Answer 3

• right-handed and asymmetrical
• complimentary base pairs (A with T and G with C)
• 10 base pairs per turn
• 0.34 nm between stacked basses and 3.4 nm per helical turn
• anti-parallel 5’ to 3’ strands

Question 4

why do AT and GC pair

Answer 4

• the space between ladders can only fit 3 rings
• A and T are able to make 2 H-bonds
• C and G are able to make 3 H-bonds

Question 5

semi-conservative copying mechanism

Answer 5

• after one round of DNA replication each DNA contains 1 parental and 1 newly-synthesized strand

Question 6

conservative replication

Answer 6

• after first replication one new and one parental strand
• after second replication one parental and 3 new strands

Question 7

dispersive replication

Answer 7

nothing is conserved
- after first replication 2 half strands
- after second replication 4 half strands

Question 8

semiconservative replication products

Answer 8

• after first replication 2 half strands
• after second replication 2 half strands and 2 new strands

Question 9

Meselson and Stahl experiment

Answer 9

used cesium chloride equilibrium-density gradient centrifugation to separate double-stranded DNA molecules of different densities
- heavier DNA sediments near bottom and lighter ones near the top
- concluded that DNA replication in E. coli is semiconservative

Question 10

Theta replication

Answer 10

• replication that occurs in most circular DNA (bacterium)
• replication is bidirectional
• unwinding at replication origin produces single-stranded templates (making a replication bubble) with replication forks at the end proceed around the circle
• eventually 2 circular DNA molecules are produced

Question 11

rolling circle replication

Answer 11

• occurs on the F factor of some viruses
• replication is unidirectional
• break in a nucleotide strand causes DNA synthesis to begin at 3’ end of the broken strand, inner strand is used as the template, 5’ broken end is displaced
• cleavage releases a single-stranded linear DNA (which may circularize) and double-stranded circular DNA
• products are multiple circular DNA’s

Question 12

Linear chromosome replication

Answer 12

• occurs in the linear chromosomes of eukaryotic cells
• replication is bidirectional
• numerous origins of replication where DNA unwinds producing a replication bubble
• DNA synthesis takes place on both strands at each end of the bubble, replication fork proceeds outward and eventually reaches another where DNA segments will fuse
• product is 2 identical linear DNA

Question 13

RNA primase

Answer 13

helps DNA polymerase
- synthesizes a bit of RNA where you want DNA polymerase to start and contains an -OH group

Question 14

DNA replication requirements

Answer 14

• magnesium (Mg2+)
• DNA dependent DNA polymerase
• 4 dNTPs
• a template DNA to be copied
• an RNA primer (provides 3’-OH end to initiate DNA synthesis by DNA polymerase)

Question 15

features of DNA replication

Answer 15

• always synthesized 5’ to 3’
• ## newly synthesized strand is complementary and anti-parallel to parent strand

Question 16

how is new DNA synthesized from dNTPs

Answer 16

• 3’-OH group of the last nucleotide on the strand attacks the 5’ phosphate of the incoming dNTP
• two phosphates are cleaved off and a phosphodiester bond forms between the 2 nucleotides
• the last incorporated nucleotide bonds with the alpha phosphate of the incoming nucleotide

Question 17

DNA chains and nuclease cleavage

Answer 17

• single and double-stranded DNA is susceptible to cleavage by nucleases
• chain cleavage can leave the alpha phosphate group attached to the 5’ or 3’ carbon of the sugar

Question 18

leading strand

Answer 18

• continuous DNA synthesis
• helices and polymerase are moving in the same direction so the polymerase doesn’t need to pause

Question 19

lagging strand

Answer 19

• discontinuous DNA synthesis
• made in segments since the polymerase needs to wait for another section of the DNA to get unwound by helicase

Question 20

DNA synthesis on leading and lagging strands

Answer 20

• DNA synthesis proceeds 5’ to 3’, same direction as unwinding
• on lagging strand, synthesis proceeds in the direction opposite of unwinding and runs out of template. it starts again, each time proceeding away from the replication fork which creates Okazaki fragments
• on the leading strand, synthesis proceeds in the same direction as unwinding making one long strand

Question 21

replication fork

Answer 21

where the helices unwinds the DNA and the polymerase follows it

Question 22

what makes the rolling circle model of replication different from the others

Answer 22

• replicates one strand at a time and doesn’t make a replication bubble
• no lagging strand because polymerase and helicase go the same way

Question 23

origin of replication: theta model

Answer 23

• DNA unwinds at the origin
• at each fork synthesis of the leading strand proceeds continuously
• synthesis of the lagging strand proceeds discontinuously

Question 24

origin of replication: rolling circle model

Answer 24

• continuous DNA synthesis begins at 3’ end of broken nucleotide strand
• as the DNA molecule unwinds, the 5’ end is progressively displaced

Question 25

origin of replication: linear eukaryotic

Answer 25

- at each fork the leading strand is synthesized continuously in the same direction of unwinding - the lagging strand is synthesized discontinuously in the direction opposite of unwinding

Question 26

proteins involved in prokaryotic DNA

Answer 26

- one 13-base pair tandem sequence (AT rich - easier to separate than GC) - four 9-base pair initiation protein binding sites

Question 27

priming of oriC

Answer 27

- initiator proteins (DnaA) bond to ori C, the origin of replication causing small stretch of the DNA to unwind - unwinding allows helicase and other single-stranded-binding proteins to attach to the single-stranded DNA

Question 28

DNA helicase in prokaryotic replication

Answer 28

- helicase unwinds the DNA 5' to 3' and travels on the lagging strand breaking H-bonds and moving the replication fork - unwound single-stranded DNA is coated with single-stranded binding protein to keep it single-stranded

Question 29

true or false: no DNA has free ends

Answer 29

true

Question 30

DNA gyrase

Answer 30

-released the stress caused by helicase by cutting one strand and letting it unwind a bit and wind back up again so it doesn't create tension as the replication bubble gets bigger

Question 31

what happens when there is more positive supercoiling

Answer 31

more resistant the strand becomes yo being unwound

Question 32

DNA primase

Answer 32

synthesizes short RNA primer that provides the 3'-OH end for DNA polymerase to begin synthesis - leading strand started first followed by lagging strand

Question 33

5 DNA polymerases in E.coli

Answer 33

1 and 3: chromosomal DHA replication 2,4 and 5: DNA repair functions

Question 34

replicative polymerases in prokaryotes

Answer 34

Pol 1: - aids removal of RNA primers - short tract synthesis - has 5' to 3' and 3' to 5'polymerase and exonuclease activity - proofreading 5' to 3' exonuclease activity Pol 3: - main replicative polymerase - highly processive (synthesizes lots of reactions without pausing) - has 5' to 3' polymerase activity - proofreading 5' to 3' exonuclease activity

Question 35

beta sliding clamp

Answer 35

a ring-shaped polypeptide that encircles the DNA and interacts with DNA polymerase 3 to enhance processive DNA synthesis - helps keep polymerase on the DNA

Question 36

lagging strand and DNA polymerase

Answer 36

when Pol3 reaches 5' end of the RNAprimer Pol3 is swapped for Pol 1 - Pol1 removes the RNA primer and resynthesizes a short tract DNA

Question 37

Elongation of DNA during replication in prokaryotes

Answer 37

-Pol 3 completes one Okazaki fragment then moves to initiate DNA synthesis of a new one - in the mean time Pol 1 exchanges places with Pol 3 to remove RNA primers - DNA must form a loop so both strands can replicate simultaneously

Question 38

All proteins involved in prokaryotic DNA replication

Answer 38

- topoisomerase (gyrase) - helicase - single-stranded binding protein - DNA primase - DNA polymerase III (and beta clamp) - DNA polymerase I - DNA ligase

Question 39

What makes eukaryotic chromosome replication unique?

Answer 39

- shorter RNA primers and Okazaki fragments - replication only occurs in the S phase -multiple polymerases (at least 15) - bidirectional replication from multiple origins on each chrom. - nucleosomes that need to be removed from parental DNA and re-assembled onto new DNA -telomeres: shorten at each round

Question 40

telomeres

Answer 40

protect the ends of chromosomes from degradation

Question 41

what is the problem with telomeres and eukaryotic DNA replication

Answer 41

- the chromosome end will be degraded causing chromosome shortening every round - we can't put a primer right at the end of DNA since telomere is there and primate needs to sit on chromosome

Question 42

how does replication happen on linear DNA with telomeres

Answer 42

- the terminal primer is positioned 70-100 nucleotides from the end of the chromosome which leaves a gap that is not replicated