12-2: DNA Replication
Process Overview

DNA “unzips” down middle

Original strands act as templates (= guides) for the new strand to build off of

Complementary bases attach  form complementary strands
o “Semi-conservative replication”---since half the original molecule is conserved in
each of the new molecules
DNA REPLICATION PROCESS

occurs in S-phase of interphase
1) Replication begins with attachment of Helicase = enzymes that break the H-bonds linking
bases, opening up the helix
o Topoisomerase = help relieve strain in remainder of helix (allowing them to swivel)
o opening proceeds in both directions, forming a replication bubble
o forms a Y-shaped structure = Replication Fork

prokaryotes  1 origin of replication

eukaryotes  Many origins of replication
2) RNA Primers are added to the DNA strand

RNA Primers = nucleotides of RNA that serve as a beginning for a new strand

RNA forms H-bonds with DNA in similar way
C≡G, but A= U
RNA Primase = enzyme that aids synthesis of RNA primers on DNA

RNA primers provide correctly paired nucleotides which have exposed 3’-OH groups
o The 3’ OH group is where the DNA nucleotide attaches
3) DNA polymerase attaches DNA nucleotides in sequence

DNA Polymerase = aid synthesis of complementary strands of DNA

Adds one nucleotide at a time to the growing strand in 5’  3’ direction
o The strand being built in this direction = Leading strand
o The other strand is backward = Lagging strand (running 3’  5’), so it must be
built in reverse in small segments = Okazaki Fragments

Very fast: adds 500-1000 nucleotides added per second

Okazaki fragments are added to until they hit the next Okazaki fragment

Leading strand goes until it hits Okazaki fragment from next replication fork
4) When 5’ end is reached:

DNA Polymerase removes RNA nucleotides and replace with DNA nucleotides

DNA Ligase connects new DNA segments to growing strand by bonding adjacent sugar
and phosphates
END REPLICATION PROBLEM

DNA polymerase can only add to the 3’ OH group of the preceding nucleotide

On the very ends, if the RNA Primer is removed without DNA nucleotides being attached
then linear DNA strands would grow successively shorter with time, deleting genes

Telomeres = repeats of short noncoding nucleotide sequences that are found at the ends of
linear (eukaryotic) chromosomes

Telomerase = enzyme that adds additional telomere nucleotides to the ends, providing a
small segment where RNA primers can be added and DNA polymerase can fill in the gap
o Prevents the shortening of linear chromosomes
o May be the “fountain of youth”, among other things (see article)
PROOFREADING

DNA polymerase only adds nucleotides if preceding nucleotides are correctly paired

If wrong, DNA polymerase reverses removing nucleotides until it reaches a correct pair, then
resumes in 5’ to 3’ direction

DNA Repair Enzymes = also “snip out” and replace wrong nucleotides
o Ex. thymine dimer “kinks”
o Error rate is 1 in 100 million base pairs
ENERGETICS OF DNA REPLICATION

Nucleotide are assembled in triphosphate form
o The P~P groups provide energy  removal of the P~P fragment make the reaction
highly exergonic
o Ensures the reaction will proceed in one direction, preventing the removal of
nucleotides
Prokaryotic Replication

Bacteria have only one single, circular chromosome to replicate

Uses only 1 origin of replication

Replication usually proceeds in both directions until they meet on the other side = Theta (Ө)
Replication

Takes 20-40 minutes
DNA AS A CARRIER OF INFORMATION

Carries from 5000 to 5 billion nucleotides

Allows for many different combinations

This explains the diversity of life on Earth