DNA synthesis Flashcards
Features of the DNA helix: importance for replication
how many bp per turn in B form DNA?
what links base pairs together?
Two antiparallel* polynucleotide chains form a RH helix, in which the bases are on the inside, phosphates and sugars on outside. 10-bp per turn in B form DNA
The polynucleotide chains are linked together by hydrogen bonds between pairs of bases- A,T and G,C. Therefore..
One strand is complementary in sequence to the other
5′…AGCTACG…3′ (*strands are antiparallel)
3′…TCGATGC…5′
Thus, copying each strand generates two daughter duplexes
DNA synthesis requires unwinding and opening of the helix, followed by copying of each DNA strand. How is it done?
What is DNA Synthesis
what does it mean by semi-conservative?
- Most human DNA has a B helix
- Errors in replication amount to 1 error/10^10 bases
• DNA replication/synthesis is semi conservative
parental DNA unwound
complementary new strand added
2 new daughter DNAs formed
How does replication begin
what is replication origin? explain what happens (3)
what recognises these areas? what do they do?
when dna unwinds at DNA origin, what forms? what does this allow?
when does this process occur in the cell cycle?
what does it involve>
length of replication for bacterial cell? mammalian cell?
- There are certain areas on the DNA called replication origins
- bacterial cell has a single replication origin
• in eukaryotes, DNA replication begins at many origins
- replication bubbles form
- bubbles move on either side
- DNA is formed
- This area is recognized by protein complexes called initiation complexes
- This initiation complex tags the replication origin
- DNA at the origin unwinds, forming a replication bubble, allowing replication machinery to enter (DNA Polymerase)
• This process occurs in the S phase of the cell cycle and involves complete unwinding of the parental DNA
- the bacterial cell cycle lasts for 20-30 mins
- mammalian cell cycle lasts for 16-24 hours
Cells have many different types of DNA Polymerases
what are the 3 types for bacteria?
what are the 5 types for eukaryotic cells?
functions?
Cells have many different types of DNA Polymerases
Bacterial cells have 3+ different types of polymerases
I: repair
II: repair
III: replication
- III was discovered as a result of mutant bacteria without I and II still showing growth
Eukaryotic cells have 5+ different types of polymerases
α: replication
- this polymerase initiates and synthesizes new DNA
β: repair
γ: mitochondrion
δ: replication
ε: replication
two main DNA polymerases are δ and ε which causes elongation
DNA polymerase key properties
acts in which direction?
what does it require? (4)
what end can you only add blocks to? what are these blocks?
what does the hydroxyl group do? effect of this?
what is the significance of DNA polymerase only acting in the 5’ - 3’ way? what does this mean? what is the leading and lagging strand? what forms on the lagging strand?
Acts in 5’ to 3’ direction, which has important consequences.
Utilises AT and CG base pairing to synthesise new DNA strands
It requires a DNA template, a DNA or RNA primer (because DNA polymerase requires an –OH on the 3’ carbon to start adding nucleotides) the four deoxyribonucleoside triphosphate (dNTP) building blocks and Mg2+ ions.
It also has a proof reading function
You can only add building blocks to the 3’ end
The dNTP’s are the building blocks for DNA
The hydroxyl group reacts with (attacks) the first phosphate and causes it to release pyrophosphate (crudely, a diphosphate). This leaves the one phosphate which then reacts with the hydroxyl forming the sugar phosphate backbone, while the bases pair.
The significance that DNA polymerase can only act in the 5’ – 3’ way, means one DNA strand must be made discontinuously, while the other is made continuously.
A replication fork will appear at the replication origin. The parental strands will separate, it is important to remember they are antiparallel. On the 5’– 3’ strand, DNA polymerase will cause a continuous strand to be made, called the leading strand, as the DNA opens up and the DNA polymerase can continue to synthesise the complementary strand.
However, on the other strand, DNA polymerase will be working in the opposite direction, this means that behind it, the strand will be opening up while it moves forward. Causing it to be called the lagging strand. This forms okazaki fragments. As replication has to be re-initiated again and again.
DNA replication involves multiple enzymes
what are the 7 enzyes?
functions of the enzymes?
Helicase = Separates the base pairs producing single strands
Topoisomerase = Goes in front of helicase and gets rid of the coils in DNA (as becomes supercoiled due to helicase) every 20 base pairs
Primase = Lays down some RNA (produces 3’ –OH), so that DNA polymerase knows where to start replicating on lagging strand
DNA binding proteins = Stabilise the single stranded DNA, stop it being re-annealed with the other parent strand
Replicative DNA polymerase = Copies the parental strand
Repair DNA polymerase = Repair the fragments and takes out the RNA so it will be a full DNA strand
DNA ligase = splices the fragments together
Proof Reading and Errors in DNA Synthesis
why is error rate low? (2)
DNA replication proceeds with high fidelity (checks for mistakes and if made will go back and change them).
Error rate is extremely low, because:
DNA polymerase error rate is about 1 in 108, due to base pairing and proof reading/editing function of the enzyme
The mismatch repair system, which corrects most of the polymerase errors. This multi-enzyme system is highly conserved across species
Defects in DNA synthesis
colon cancer? how?
how can they be used in treatment?
Inherited defects in mismatch repair genes are involved in (colon) cancer, in essence, failure to correct DNA synthesis may lead to cancer
DNA replication inhibitors are important antibacterial, antitumour (so used in chemotherapy against cancers) and antiviral agents.
