Mod 2 - DNA replication Flashcards
What is DNA replication? (semi-conservative)
when two DNA strands seperate and DNA is copied using the parent strand as a template
What did the meselson-stahl experiment set out to do?
to ascertain what type of DNA replication occurs in e.coli bacteria (between semiconservative, conservative, and dispersive)
performed via growing bacteria in a heavy nitrogen (15N) medium, then distinguishing the 14N bacteria from the 15N bacteria via density gradient centrifugation
what is the mode of action of type 1 DNA topoisomerase?
makes a single-strand nick to prevent supercoiling
what is the mode of action of type 2 DNA topoisomerase?
makes a double strand cut to prevent supercoiling
in what direction does DNA replication occur?
5’ -> 3’
what is the purpose of DNA helicase?
to break the hydrogen bonds between the base pairs to seperate the DNA strands
what is 3’ -> 5’ exonuclease activity?
the mechanism by which polymerase can remove nucleotides that it has just inserted (proofreading)
what is 5’ -> 3’ exonuclease activity?
the mechanism by which polymerase can remove DNA that is already attatched to the template
what are the 2 bacterial DNA polymerases?
DNA Pol 1 and DNA Pol 3 (III)
what are the 2 eukaryotic DNA polymerases?
DNA polymerase alpha (α) and DNA polymerase delta (δ)
what are some features of DNA Pol I ? (4)
- 1 subunit
- can do both 3’ ->5’ AND 5’ -> 3’ exonuclease activity
- used for DNA repair and replication
- acts as a DNA-dependent DNA polymerase
what are some features of DNA Pol III ? (4)
- at least 10 subunits
- can do 3’ -> 5’
- can’t do 5’ -> 3’
- acts as main replicating enzyme
main replicating enzyme therefore can’t do 5’ -> 3’ exonuclease activity
what are some features of DNA Pol alpha ? (3)
- 4 subunits
- can’t do 3’ -> 5’ OR 5’ -> 3’
- used for priming during replication
basically fucking useless. very bad alpha
what are some features of DNA Pol delta ? (4)
- 2 or 3 subunits
- can do 3’ -> 5’
- can’t do 5’ -> 3’
- acts as main replicating enzyme
main replicating enzyme therefore can’t do 5’ -> 3’ exonuclease activity
what protects the single strands at the replication fork from re-attatching?
single-strand binding proteins (SSB’s)
how does the primer work in bacteria?
- made of primase enzyme
- primer is normally 4-15 nucleotides long
- once primer is made, DNA Pol III makes new strand
how does the primer work in eukaryotes?
- RNA primer extended by DNA Pol alpha by 20 nucleotides
- DNA Pol delta makes the rest of the new strand
where is the primer placed for replication of the lagging strand?
at the replication fork - DNA must locate in a 5’ to 3’ direction
how is the lagging strand replicated?
In sections, as when the replication fork moves along the strand, the primer must also move along with it
- forms okazaki fragments when primers are removed
how are okazaki fragments joined in bacteria?
- DNA Pol III synthesises DNA until it reaches primer
- DNA Pol I continues synthesis after primer
- DNA ligase joins fragments together
how are okazaki fragments joinedi in eukaryotes?
-DNA Pol delta continues to synthesise DNA through the primer -
with the aid of DNA helicase, it pushes the primer to one side, creating a branch point
- FEN1 cuts at the branch point
- DNA ligase joins fragments together
what is the end replication problem?
the final okazaki fragment cannot be made as the priming site would be after the end of the parent molecule
- this problem operates on the basis that the primer must attatch to bases.
- would, over time, cause the molecule to become shorter
what is the solution to the end replication problem?
telomerase
- extends parental DNA by adding the sequence TTAGGG several times
- adds DNA that can be sacrificed with no adverse affects, as it was not part of the original parent molecule.
what kind of cells have telomerase?
stem cells
what is senescence?
regular cells dying due to the ends of the chromosome shortening as they do not have telomerase to overcome the end replication problem
