The two replication forks move in opposite directions

The two replication forks move in opposite directions sequentially around the circular
chromosome
around(the(circular(chromosome(
unique ori !
fully !
replicated !
The Phage  Chromosome
 Three forms of  DNA
o Linear
o H-bonded
o Covalently bonded circular
 AT-rich and GC-rich regions used to
demonstrate bidirectional replication
 Denaturation mapping
 Increase temperature or pH to denature
genome
 Easier to break A-T bonds than G-C
bonds because G-C have more hydrogen
bonds
By examining the positions of the branch point
(Y-shaped structures) relative to the positions
of the denaturing bubbles in a large number of
Y-shaped replicative intermediates, Schnos and
Inman demonstrated that both branch points
are replication forks that move in opposite
directions around the circular chromosome
Key Points
 DNA replicates by a semiconservative
mechanism: as the two complementary
strands of a parental double helix

unwind and separate, each serves as template for the synthesis of a new complementary
strand
 The hydrogen bonding potentials of the bases in template strands specify
complementary base sequences in the nascent DNA strands
 Replication is initiated at unique origins and usually proceeds bidirectionally from each
origin
Exercise:
 The Escherichia coli chromosome contains approx. 4 x 10 6 nucleotide pairs and replicates
as a single bidirectional replicon in approx. 40 minutes under a wide variety of growth
conditions.
 The Supermegabigia coli chromosome contains approx. 8 x 10 7 nucleotide pairs and
replicates at the same rate as E.coli
 How long does it take to replicate the Supermeganigia coli?
Is the answer:!
!
a) 80 minutes!
!
b) 120 minutes!
!
c) 800 minutes!
!
d) 2000 minutes!
Because each chromosome replicates bidirectionally,
each replication fork must traverse 2 x 10
6
nucleotide
pairs in E. coli (or half the total nucleotide pairs).!
Same principle applies to “ Supermegabigia# coli ”
chromosome; it must traverse 4 x 10
7
nucleotide pairs.!
4 x 10
7
nucleotide pairs in “ Supermegabigia# coli ” !
2 x 10
6
nucleotide pairs in E. coli !
!
= 20 times more nucleotide pairs in !
“ Supermegabigia# coli ” !
40 minutes to replicate
all the nucleotide pairs in
E. coli x 20 !
!
= 800 minutes !
!
DNA Replication in Prokaryotes
 Must understand the process and the different proteins that enable this process
 Names of proteins involved
 How the process works
 Know how each protein works
 Don’t need to know structures of proteins
 Won’t be asked how many subunits

Many different DNA polymerases
 E.coli contains at least 5 different DNA polymerases:
o DNA polymerase I –multiple functions (polymerase and nuclease)
o DNA polymerase II – replication of damaged DNA
o DNA polymerase III – multiple functions (polymerase and nuclease)
o DNA polymerase IV – replication of damaged DNA
o DNA polymerase V – replication of damaged DNA
(dNMP)
n
+ dNTP (dNMP)
n+1
+ PP
i
DNA template
DNA dependent
DNA polymerase
 DNA replication cannot occur de novo (from nothing)
o Primer with a free OH group at its 3’ end
o Template DNA to specify the sequence of the new DNA strand