Download Ch.16 DNA Structure & Replication notes

Evidence that DNA can transform bacteria
• Frederick Griffith (1928) – Streptococcus
pneumoniae bacteria – transformation
• Mouse Experiment
• Experiment proved that transformation can
happen
• Avery and colleagues (1944) – announced
transformation agent was DNA
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Figure 16.2 Can the genetic trait of pathogenicity
be transferred between bacteria?
EXPERIMENT Bacteria of the “S” (smooth) strain of Streptococcus pneumoniae are pathogenic because they
have a capsule that protects them from an animal’s defense system. Bacteria of the “R” (rough) strain lack a capsule
and are nonpathogenic. Frederick Griffith injected mice with the two strains as shown below:
Living S
(control) cells
Living R
Heat-killed
(control) cells (control) S cells
Mixture of heat-killed S cells
and living R cells
RESULTS
Mouse dies
Mouse healthy
Mouse healthy
Mouse dies
Living S cells
are found in
blood sample.
CONCLUSION Griffith concluded that the living R bacteria had been transformed into pathogenic S bacteria by an
unknown, heritable substance from the dead S cells.
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Evidence that viral DNA can program cells
• Alfred Hershey and Martha Chase (1952) –
bacteriophages or phages (viruses that infect
bacteria) – discovered DNA is the genetic
material NOT protein
• Blender experiment
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Figure 16.3 Viruses infecting a bacterial cell
Phage
head
Tail
Tail fiber
Bacterial
cell
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100 nm
DNA
Additional evidence that DNA is the genetic
material
• Erwin Chargaff (1947) – Chargaff’s rules – The
equivalences for any given species between
the number of A and T and G and C bases are
equal.
• Analyzed DNA from different organisms
– Humans 30.3% of bases were A’s
– E. Coli 26% of bases were A’s
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• Rosalind Franklin – (1950’s) – X-ray diffraction
photo of DNA – helped Watson and Crick
develop their model of DNA structure
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Figure 16.6 Rosalind Franklin and her X-ray
diffraction photo of DNA
(a) Rosalind Franklin
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(b) Franklin’s X-ray diffraction
Photograph of DNA
Structure of DNA
• Watson & Crick – (1953) – 1 page paper in the
British journal Nature “Molecular Structure of
Nucleic Acids: A Structure for Deoxynucleic
acids”
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Figure 16.1 Watson and Crick with their DNA model
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Figure 16.5 The structure of a DNA strand
Sugar-phosphate
backbone
5 end
5
O P O CH2
Nitrogenous
bases
CH3
O–
O–
4 H
H
3
O
H
O
1
N
N
H
H
O
Thymine (T)
H
2
H
O
O
P O
CH2
O–
H
H
N
O
H
H
H
H
Adenine (A)
H
H
O
P O
H
N
N
H
O
N
N
H
CH2
O–
H
O
H
N H
N
H
N
H
H
O
Cytosine (C)
H
O
O
5
P O CH2
H
O
1
O– 4 H
H
Phosphate H
H
2
3
H
OH
Sugar (deoxyribose)
3 end
N
O
N
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N
N H
N H
H
Guanine (G)
DNA nucleotide
Figure 16.7 The double helix
G
5 end
C
O
A
T
–O
T
A
P
OH
O
H2C
Hydrogen bond
O
O
C
C
3.4 nm
G
–O
P
O
O
G
T
A
–O
A
T
O
CH2
G
O
P
O
O
H2C
O
O
A
T
P
O–
O
T
A
OH
3 end
G
C
0.34 nm
A
CH2
O
O–
P
O
O
O
C
O
O
O
O
O–
P
C
O
H2C
A
CH2
O
O
H2C
O
G
T
O
O
T
A
C
–O
P
A
T
1 nm
G
3 end
OH
T
(a) Key features of DNA structure
O
CH2
O
O–
P
O
O
5 end
(b) Partial chemical structure
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(c) Space-filling model
Figure 16.8 Base pairing in DNA
H
N
N
N
N
Sugar
O
H
H
CH3
N
N
N
O
Sugar
Thymine (T)
Adenine (A)
H
O
N
N
Sugar
N
H
H
N
N
N
N
N
H
Guanine (G)
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H
O
Sugar
Cytosine (C)
Unnumbered Figure p. 298
Purine + Purine: too wide
Pyrimidine + pyrimidine: too narrow
Purine + pyrimidine: width
Consistent with X-ray data
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DNA Replication Section 16.2
• Semi-conservative model – each of the two
daughter molecules will have one old strand,
derived from the parent molecules, and one
newly made strand
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Figure 16.9 A model for DNA replication: the basic
concept (layer 1)
A
T
C
G
T
A
A
T
G
C
(a) The parent molecule has two
complementary strands of DNA.
Each base is paired by hydrogen
bonding with its specific partner,
A with T and G with C.
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Figure 16.9 A model for DNA replication: the basic
concept (layer 2)
A
T
A
T
C
G
C
G
T
A
T
A
A
T
A
T
G
C
G
C
(a) The parent molecule has two
complementary strands of DNA.
Each base is paired by hydrogen
bonding with its specific partner,
A with T and G with C.
(b) The first step in replication is
separation of the two DNA
strands.
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Figure 16.9 A model for DNA replication: the basic
concept (layer 3)
T
A
T
A
T
A
C
G
C
G
C
T
A
T
A
T
A
A
T
A
T
A
T
G
C
G
C
G
C
G
A
T
C
G
T
A
A
C
G
(a) The parent molecule has two
complementary strands of DNA.
Each base is paired by hydrogen
bonding with its specific partner,
A with T and G with C.
(b) The first step in replication is
separation of the two DNA
strands.
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T
(c) Each parental strand now
serves as a template that
determines the order of
nucleotides along a new,
complementary strand.
Figure 16.9 A model for DNA replication: the basic
concept (layer 4)
T
A
T
A
T
A
C
G
C
G
C
T
A
T
A
T
A
A
T
A
T
A
T
G
C
G
C
G
C
G
A
T
A
T
A
T
C
G
C
G
C
G
T
A
T
A
T
A
T
A
T
A
T
C
G
C
G
C
A
G
(a) The parent molecule has two
complementary strands of DNA.
Each base is paired by hydrogen
bonding with its specific partner,
A with T and G with C.
(b) The first step in replication is
separation of the two DNA
strands.
Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
(c) Each parental strand now
serves as a template that
determines the order of
nucleotides along a new,
complementary strand.
(d) The nucleotides are connected
to form the sugar-phosphate
backbones of the new strands.
Each “daughter” DNA
molecule consists of one parental
strand and one new strand.
Figure 16.10 Three alternative models of DNA replication
Parent cell
(a) Conservative
model. The two
parental strands
reassociate
after acting as
templates for
new strands,
thus restoring
the parental
double helix.
(b) Semiconservative model.
The two strands
of the parental
molecule
separate,
and each functions
as a template
for synthesis of
a new, complementary strand.
(c) Dispersive
model. Each
strand of both
daughter molecules contains
a mixture of
old and newly
synthesized
DNA.
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First
replication
Second
replication
Figure 16.13 Incorporation of a nucleotide into a DNA strand
New strand
Template strand
Sugar
A
Base
Phosphate
3’ end
5’ end
3’ end
5’ end
T
A
T
C
G
C
G
G
C
G
C
A
T
A
P
OH
P
Pyrophosphate 3’ end
C
C
OH
Nucleoside
triphosphate
2 P
5’ end
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5’ end
Figure 16.12 Origins of replication in eukaryotes
Origin of replication
1 Replication begins at specific sites
where the two parental strands
separate and form replication
bubbles.
Bubble
Parental (template) strand
Daughter (new) strand
0.25 µm
Replication fork
2 The bubbles expand laterally, as
DNA replication proceeds in both
directions.
3 Eventually, the replication
bubbles fuse, and synthesis of
the daughter strands is
complete.
Two daughter DNA molecules
(a) In eukaryotes, DNA replication begins at many sites along the giant
DNA molecule of each chromosome.
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(b) In this micrograph, three replication
bubbles are visible along the DNA of
a cultured Chinese hamster cell (TEM).
Figure 16.14 Synthesis of leading and lagging
strands during DNA replication
1 DNA pol Ill elongates
DNA strands only in the
5
3 direction.
3
5
Parental DNA
2 One new strand, the leading strand,
can elongate continuously 5
3
as the replication fork progresses.
5
3
Okazaki
fragments
2
1
3
5
3 The other new strand, the
lagging strand must grow in an overall
3
5 direction by addition of short
segments, Okazaki fragments, that grow
5
3 (numbered here in the order
they were made).
DNA pol III
Template
strand
3
Leading strand
Lagging strand
2
Template
strand
1
DNA ligase
Overall direction of replication
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4 DNA ligase joins Okazaki
fragments by forming a bond between
their free ends. This results in a
continuous strand.
Figure 16.15 Synthesis of the lagging strand
1
Primase joins RNA nucleotides
into a primer.
3
5
5
3
Template
strand
2
RNA primer
3
5
3
DNA pol III adds DNA nucleotides to the
primer, forming an Okazaki fragment.
5
3
1
After reaching the next
RNA primer (not shown),
DNA pol III falls off.
Okazaki
fragment
3
3
5
1
5
4
After the second fragment is
primed. DNA pol III adds DNA
nucleotides until it reaches the
first primer and falls off. 5
3
5
3
2
5
1
DNA pol 1 replaces the
RNA with DNA, adding to
the 3 end of fragment 2.
5
3
6
3
2
DNA ligase forms a bond
between the newest DNA
and the adjacent DNA of
fragment 1.
5
3
5
1
7
The lagging strand
in this region is now
complete.
3
2
1
Overall direction of replication
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5
Table 16.1 Bacterial DNA replication proteins and
their functions
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Figure 16.16 A summary of bacterial DNA replication
Overall direction of replication
Lagging
Leading
strand Origin of replication strand
1 Helicase unwinds the
parental double helix.
2 Molecules of single- 3 The leading strand is
strand binding protein synthesized continuously in the
stabilize the unwound 5 3 direction by DNA pol III.
template strands.
DNA pol III
Lagging
strand
OVERVIEW
Leading
strand
Leading
strand
5
3
Parental DNA
4 Primase begins synthesis
of RNA primer for fifth
Okazaki fragment.
5 DNA pol III is completing synthesis of
the fourth fragment, when it reaches the
RNA primer on the third fragment, it will
dissociate, move to the replication fork,
and add DNA nucleotides to the 3 end
of the fifth fragment primer.
Replication fork
Primase
DNA pol III
Primer
4
Lagging
strand
3
6 DNA pol I removes the primer from the 5 end
of the second fragment, replacing it with DNA
nucleotides that it adds one by one to the 3’ end
of the third fragment. The replacement of the
last RNA nucleotide with DNA leaves the sugarphosphate backbone with a free 3 end.
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DNA ligase
DNA pol I
2
1
3
5
7 DNA ligase bonds
the 3 end of the
second fragment to
the 5 end of the first
fragment.
Proofreading and repairing DNA
• Errors do occur
– 1 in 10 billion nucleotides on entire DNA
– 1 in 100,000 for incoming nucleotides
• Proofreading is done by DNA pol III as it
attaches new nucleotides
• Mismatch repair cells use special enzymes to
fix mismatched nucleotides
– A-C for example
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Figure 16.17 Nucleotide excision repair of DNA damage
1 A thymine dimer
distorts the DNA molecule.
2 A nuclease enzyme cuts
the damaged DNA strand
at two points and the
damaged section is
removed.
Nuclease
3 Repair synthesis by
a DNA polymerase
fills in the missing
nucleotides.
DNA
polymerase
DNA
ligase
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4 DNA ligase seals the
Free end of the new DNA
To the old DNA, making the
strand complete.
Telomeres
• Repeated units of bases
– TTAGGG in humans
• Do NOT contain genes
• They protect the genes from being eroded
(getting shorter and shorter) through DNA
replication rounds
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Figure 16.18 Shortening of the ends of linear DNA molecules
5
End of parental
DNA strands
Leading strand
Lagging strand
3
Last fragment
Previous fragment
RNA primer
Lagging strand
5
3
Primer removed but
cannot be replaced
with DNA because
no 3 end available
for DNA polymerase
Removal of primers and
replacement with DNA
where a 3 end is available
5
3
Second round
of replication
5
New leading strand 3
New lagging strand 5
3
Further rounds
of replication
Shorter and shorter
daughter molecules
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Figure 16.19 Telomeres
1 µm
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