DNA replication Flashcards
What type of replication does DNA undergo?
Semi Conservative
What is required to form a new DNA strand?
Template strand, free DNA nucleotides (dNTPs), enzymes
What is the pre-replication complex?
Protein complex that forms at the origin of replication
What is required to initiate replication?
Formation of the pre-RC
What enzymes promote DNA opening ?
DNA helicase
How does DNA helicase promote DNA opening?
Cleaving the inter-base hydrogen bonds using ATP hydrolysis energy.
What enzyme synthesises short RNA primers?
Primase
What do the RNA primers do?
Primers anneal to the strand of DNA to initiate replication and the action of DNA polymerase.
Which strand are RNA primers added to?
RNA primer added to both the leading and lagging strands.
Why is a primer used?
Provides a 3-OH’ (prime) terminus to initiate replication
How many primers are needed in the leading strand vs lagging strand?
1 in leading (at the start)
Several in lagging for each okazaki fragment
What about DNA means there is a lagging and a leading strand?
Anti-parallel
DNA polymerase can only add new dNTPs to the … end
3’
DNA polymerase can only add new dNTPs to the 3’ end in a ….. direction
5’ to 3’
Where does the DNA polymerase move in the leading strand?
5’ to 3’ (following new strand)
How does DNA polymerase move along the strand?
Sliding clamp - PCNA
What direction does the template strand lie in leading strand?
3’-5’
What direction does the template strand lie in lagging strand?
5’-3’
Why cant DNA polymerase not add dNTPs on lagging strand in 3’-5’ direction?
DNA polymerase can only add to 3’ end
What direction does the new strand lie in leading strand?
5’-3’
What direction does the new strand lie in lagging strand?
3’-5’
How can both strands be replicated in the same direction?
Okazaki fragments
Lagging strand replicated in ‘backstitch’ method
Describe how the lagging strand is replicated
Primase inserts a primer onto the DNA in the direction the DNA needs to grow
Polymerase associates with the primer and will move 5’ to 3’ away from the primer in the opposite direction as the leading strand.
Then another primer is placed further back until the full strand is replicated in Okazaki fragments.
What is the function of PCNA?
Clamp that tethers polymerase to the DNA and displaces the primase enzyme.
What proteins loads clamp onto the DNA?
Protein RFC
What does protein RFC require to attach PCNA to DNA?
ATP
What unwinds the DNA further along to prevent coil strain and risk of breaking?
Topoisomerase
What is the role of topoisomerase?
Topoisomerases unwind the DNA at fork to prevent tortional strain and risk of breaking.
What removes and degrades the RNA primer flaps in the lagging strands?
FEN1
What pushes the RNA primers out the way?
DNA polymerase
What does FEN1 do?
Removes the RNA primer flap after it is pushed aside by DNA polymerase. It also degrades it.
What joins together Okazaki fragments?
DNA ligase
What do telomeres consist of?
Long tandem repeats, constitutive chromatin
What do telomeres do?
Specific structures at the end of chromosomes that prevent chromosome shortening during cell division
Why do telomeres shorten with each division?
There is no template for primase on the telomere so RNA primer can not form for the last Okazaki fragment. So with each round there is shortening and senescence.
How is telomere shortening prevented?
Telomerase enzyme
Has an RNA component and is a reverse transcriptase so uses RNA template to make complimentary DNA.
What is a proof-reading polymerase?
Polymerases with 3’ to 5’ exonuclease activity can correct mistakes.
What can 3’-5’ exonuclease polymerases do?
Remove the wrong nucleotide when a mistake is made in replication
