Chapter 6 Flashcards
(38 cards)
What is DNA replication?
The complete, faithful copying of the DNA comprising the cell’s chromosomes.
Why is DNA replication semi-conservative?
Each strand of the parental double helix acts as a template for synthesis of a new daughter DNA strand.
Semi-conservative vs Conservative?
Semi: each new DNA molecule consists of one original strand and one newly synthesized strand
Cons: one DNA molecule remains entirely original and the other is entirely new
What should be in order to ensure identical copying? (2)
- base on template strand must be identified
- complementary bases must be added
Where does replication start? And what follows? Describe this process.
- At special sites called “origins of replication” then moves away in both directions forming a “replication bubble”
- DNA double-helix opens at origin of replication and unwinds to form “replication forks”
What happens at replication forks?
- single stranded DNA is exposed
- DNA synthesis can occur (5’ to 3’ direction)
Describe the synthesis coninuity.
Continuous on leading strand and discontinuous on lagging strand.
What are the 3 phases of synthesis?
Initiation
Elongation
Termination
Describe the initiation process.
- Initiator protein regognizes the origin of replication, then opens up the double helix
- And recruits helicases that unwinds the helix to expose single-stranded DNA (coated by ssDNA binding proteins that prevents reannealing)
How many times does initiation occur in a cycle?
Once
Why does DNA synthesis need a primer?
Can only add nucleotides to a pre-existing 3’ end - primer can initiate a new RNA strand without a pre-existing 3’ end.
How is primase used?
Primer synthesis by the primase.
The RNA primer is degraded and replaced with DNA to complete the replication.
Describe the process of elongation.
- After the RNA primer is synthesized, DNA polymerase is recruited.
- DNA polmerase is associated with DNA via the clamp loader and sliding clamp.
- The replication machinery moves along the DNA, copying the strands.
- DNA reads ech base in the parental DNA, wich adds complementary bases to he growing strand in a 5’ to 3’ direction.
Describe the process of termination.
- When two different forks meet or when the fork reaches the endof a linear chromosome.
- Replication complexes are disassembled.
- RNA primers are removed and replaced with DNA.
- DNA ligase connects adjacent strands.
The functions of DNA polymerases? (2)
- Catalyze the addition of nucleotides to the 3’ OH of the last nucleotide in the newly growing strand.
- Exonucleases remove the incorrect nucleotide if an error is found. (PROOFREAD)
What are some of the polymerases for?
- genome relication
- repair and replication across damaged DNA (translesion)
- telomere replication
What are the main polymerases involved in DNA replication?
- DNA polymerase III in bacteria
- DNA polymerase δ and ε (replicative) in eukaryotes
Describe the structure of these major polymerases.
- Highly conserved and has multiple subunits with specific functions
- Right hand with three domains
What is the processivity of a polymerase?
It remains attached to DNA for long stretches before dissociation.
Describe the 3 domains of DNA polymerase.
Palm - single-stranded DNA is fed past fingers into the active site (palm)
Finger - positions the incoming nucleotide, which joins to the free 3’OH in the polymerase active site
Thumb - holds elongating dsDNS
What happens in the active site of DNA polymerase (thumb)?
Has carboxylate groups of two aspartate resides coordinated with two magnesium ions.
It catalyzes a phosphoryl transfer reaction linking the 5’ phosphoryl group of the incoming nucleotide to the 3’ OH of the growing DNA to form a PHOSPHODIESTER BOND.
Describe the phosphoryl transfer reaction.
Consists of a nucleophilic attack by the 3’ OH (nucleophile) on the α-phosphate (electrophile) of the incoming nucleoside triphosphate.
Which releases two of the phosphates as pyrophosphate -> hydrolysis to provide free energy that drives reaction forward.
Why is the two magnesium ions at the active site critical?
- One activates the 3’OH
- The other interacts with the incoming nucleotide and stabilizes the negative leaving oxygen
Bacterial vs eukaryotic DNA polymerase?
- overall structure & catalytic mechanism of polymerases are similar
- the special relationships b/w domains differ among different polymerases
- structural differences in the catalytic palm domain = replication machinery arose x2 during evolution & convergent evolution led to similarities
