1.5 - DNA replication Flashcards
direction of DNA synthesis
5’-3’
3 possible principles of DNA replication
- semiconservative
- dispersive
- conservative
how is a primer located
ends with OH group
dNTP
deoxynucleotide triphosphate
- fundamental building block of DNA
components of deoxynucleotide triphosphate (3)
- nitrogenous base
- deoxyribose sugar
- triphosphate group
triphosphate group in dNTP
3 phosphates attached to 5’ carbon of deoxyribose sugar, provides energy for DNA polymerisation
DNA synthesis (2)
- dNTP base pairs with the template strand through H-bonding
- DNA polymerase catalyses formation of a new phospho-diester bond between the 3’OH of the preceding nucleotide in the chain with the new base pair nucleotide
rate of incorporation of incorrect dNTP
10,000-fold slower
rate of incorporation of rNTP
1000-fold slower
“discriminator” amino acids
in DNA polymerase, prevent rNTP binding via hydroxyl group on discriminator amino acid creating clash with rNTP (preventing binding)
rNTP
primary substrate for RNA synthesis
how are RNA and DNA polymerases different
unlike RNA polymerase, DNA polymerase cannot initiate a new DNA chain
what is each new DNA chain initiated by
synthesis of a short RNA chain (RNA primer catalysed by a primase) extended by DNA polymerase
RNA primer
synthesised short RNA chain that initiates new DNA chain, catalyses by primase and extended by DNA polymerase
what happens to RNA primer when new DNA chain is initiated
RNA primer part of chain removed
different characteristics of DNA polymerases in eukaryotic cells (2)
- some focused on bulk DNA replication at replication fork
- some focused on bypass of damaged DNA or DNA repair pathways
how many bp does DNA polymerase synthesise before releasing template?
20-100bp
how is DNA polymerase prevented from falling off?
sliding clamp encircles newly synthesised dsDNA and holds DNA polymerase (preventing diffusion away from primer)
post replication error rate
1 mistake per 10^10 nt added
DNA polymerase insertion error rate
1 incorrect dNTP per 10^5 nt added
(base pair accuracy alone not sufficient to account for accuracy - 1 - 10^10)
how is DNA proofread? (2)
- DNA polymerases that read for DNA replication have an exonuclease site next to active site
- incorrectly incorporated bases can be caught by exonuclease activity (allowing DNA polymerase another go at incorporating correct base)
why is DNA synthesis always 5’-3’?
proofreading is chemically difficult if synthesis occurred in 3’-5’ direction (proofreading critical for high fidelity of action by DNA polymerases)
why can only 1 strand be continuously synthesised at a time
only one of 2 strands can be continuously synthesised (because of 5’-3’ direction)
okazaki fragments
short DNA nucleotide sequences synthesised discontinuously on lagging strand, later joined by DNA ligase to form continuous DNA strand
