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11
DNA and Its Role in Heredity
11 DNA and Its Role in Heredity
• 11.1 What Is the Evidence that the
Gene Is DNA?
• 11.2 What Is the Structure of DNA?
• 11.3 How Is DNA Replicated?
• 11.4 How Are Errors in DNA Repaired?
• 11.5 What Are Some Applications of Our
Knowledge of DNA Structure and
Replication?
11.1 What Is the Evidence that the Gene Is DNA?
By the 1920s, it was known that
chromosomes consisted of DNA and
proteins.
A new dye stained DNA and provided
circumstantial evidence that DNA was the
genetic material:
It was in the right place
It varied among species
It was present in the right amount
11.1 What Is the Evidence that the Gene Is DNA?
Frederick Griffith, working with two
strains of Streptococcus pneumoniae
determined that a “transforming
principle” from dead cells of one strain
produced a heritable change in the
other strain.
Figure 11.1 Genetic Transformation of Nonvirulent Pneumococci
11.1 What Is the Evidence that the Gene Is DNA?
Identifying the transforming principle,
Oswald Avery:
Treated samples to destroy different
molecules; if DNA was destroyed, the
transforming principle was lost.
Figure 11.2 Genetic Transformation by DNA (Part 1)
Figure 11.2 Genetic Transformation by DNA (Part 2)
11.1 What Is the Evidence that the Gene Is DNA?
Hershey-Chase experiment:
• Determined whether DNA or protein is
the genetic material using
bacteriophage T2 virus.
• Bacteriophage proteins were labeled
with 35S; the DNA was labeled with 32P.
Figure 11.3 Bacteriophage T2: Reproduction Cycle
Figure 11.4 The Hershey–Chase Experiment (Part 1)
Figure 11.4 The Hershey–Chase Experiment (Part 2)
11.1 What Is the Evidence that the Gene Is DNA?
Next, genetic transformation of
eukaryotic cells was demonstrated—
called transfection.
Use a genetic marker—a gene that
confers an observable phenotype.
Any cell can be transfected, even an egg
cell—results in a transgenic organism.
Figure 11.5 Transfection in Eukaryotic Cells
11.2 What Is the Structure of DNA?
The structure of DNA was determined
using many lines of evidence.
One crucial piece came from X-ray
crystallography.
A purified substance can be made to
form crystals; position of atoms is
inferred by the pattern of diffraction of
X-rays passed through it.
Figure 11.6 X-Ray Crystallography Helped Reveal the Structure of DNA
11.2 What Is the Structure of DNA?
Chemical composition also provided
clues:
DNA is a polymer of nucleotides:
deoxyribose, a phosphate group, and a
nitrogen-containing base.
The bases:
• Purines: adenine (A), guanine (G)
• Pyrimidines: cytosine (C), thymine (T)
Figure 3.23 Nucleotides Have Three Components
repeat fig 3.23 here
11.2 What Is the Structure of DNA?
1950: Erwin Chargaff found in the DNA
from many different species:
amount of A = amount of T
amount of C = amount of G
Or, the abundance of purines = the
abundance of pyrimidines—Chargaff’s
rule.
Figure 11.7 Chargaff’s Rule
11.2 What Is the Structure of DNA?
Model building started by Linus Pauling—
building 3-D models of possible
molecular structures.
Francis Crick and James Watson used
model building and combined all the
knowledge of DNA to determine its
structure.
Figure 11.8 DNA Is a Double Helix (A)
11.2 What Is the Structure of DNA?
X-ray crystallography convinced them the
molecule was helical.
Other evidence suggested there were
two polynucleotide chains that ran in
opposite directions—antiparallel.
1953—Watson and Crick established the
general structure of DNA.
Figure 11.8 DNA Is a Double Helix (B)
11.2 What Is the Structure of DNA?
Key features of DNA:
• A double-stranded helix, uniform diameter
• It is right-handed
• It is antiparallel
• Outer edges of nitrogenous bases are
exposed in the major and minor grooves
11.2 What Is the Structure of DNA?
Complementary base pairing:
• Adenine pairs with thymine by two
hydrogen bonds.
• Cytosine pairs with guanine by three
hydrogen bonds.
• Every base pair consists of one purine
and one pyrimidine.
Figure 11.9 Base Pairing in DNA Is Complementary (Part 1)
Figure 11.9 Base Pairing in DNA Is Complementary (Part 2)
11.2 What Is the Structure of DNA?
Antiparallel strands: direction of strand
is determined by the sugar–phosphate
bonds.
Phosphate groups connect to the 3′ C of
one sugar, and the 5′ C of the next
sugar.
At one end of the chain—a free 5′
phosphate group; at the other end a
free 3′ hydroxyl.
11.2 What Is the Structure of DNA?
The flat base pairs are exposed in the
major and minor grooves—accessible
for hydrogen bonding.
The C═O group in thymine, the N group
in adenine, and others offer hydrogen
bonding sites.
Key to DNA–protein interactions in
replication and gene expression.
11.2 What Is the Structure of DNA?
Functions of DNA:
• Store genetic material—millions of
nucleotides; base sequence stores and
encodes huge amounts of information
• Susceptible to mutation—change in
information
11.2 What Is the Structure of DNA?
• Genetic material is precisely replicated
in cell division—by complementary base
pairing.
• Genetic material is expressed as the
phenotype—nucleotide sequence
determines sequence of amino acids in
proteins.
11.3 How Is DNA Replicated?
Kornberg showed that DNA contains
information for its own replication.
In a test tube: DNA, the four
deoxyribonucleoside triphosphates, and
DNA polymerase enzyme.
The DNA is a template for synthesis of
new DNA.
11.3 How Is DNA Replicated?
Three possible replication patterns:
• Semiconservative replication
• Conservative replication
• Dispersive replication
Figure 11.10 Three Models for DNA Replication
11.3 How Is DNA Replicated?
Meselson and Stahl showed that
semiconservative replication was the
correct model.
They used density labeling to distinguish
parent DNA strands from new DNA
strands.
DNA was labeled with 15N, making it
more dense.
Figure 11.11 The Meselson–Stahl Experiment (Part 1)
Figure 11.11 The Meselson–Stahl Experiment (Part 2)
11.3 How Is DNA Replicated?
Results of their experiment can only be
explained by the semiconservative model.
If it was conservative, the first generation of
individuals would have all been high or low
density, but not intermediate.
If dispersive, density in the first generation
would be half, but this density would not
appear in subsequent generations.
11.3 How Is DNA Replicated?
Two steps in DNA replication:
• The double helix is unwound, making
two template strands.
• New nucleotides are added to the new
strand at the 3′ end; joined by
phosphodiester linkages. Sequence is
determined by complementary base
pairing.
Figure 11.12 Each New DNA Strand Grows from Its 5′ End to Its 3′ End (Part 1)
Figure 11.12 Each New DNA Strand Grows from Its 5′ End to Its 3′ End (Part 2)
11.3 How Is DNA Replicated?
A large protein complex—the replication
complex—catalyzes the reactions of
replication.
All chromosomes have a base sequence
called origin of replication (ori).
Replication complex binds to ori at start.
DNA replicates in both directions, forming
two replication forks.
Figure 11.13 Two Views of DNA Replication
11.3 How Is DNA Replicated?
DNA helicase uses energy from ATP
hydrolysis to unwind the DNA.
Single-strand binding proteins keep
the strands from getting back together.
11.3 How Is DNA Replicated?
Small, circular chromosomes have a
single origin of replication.
As DNA moves through the replication
complex, two interlocking circular
chromosomes are formed.
DNA topoisomerase separates the two
chromosomes.
Figure 11.14 Replication in Small Circular and Large Linear Chromosomes (A)
11.3 How Is DNA Replicated?
Large linear chromosomes have many
origins of replication.
DNA is replicated simultaneously at the
origins.
Figure 11.14 Replication in Small Circular and Large Linear Chromosomes (B)
11.3 How Is DNA Replicated?
DNA polymerases are much larger than
their substrates.
Shape is like a hand; the “finger” regions
have precise shapes that recognize the
shapes of the nucleotide bases.
Figure 11.15 DNA Polymerase Binds to the Template Strand (Part 1)
Figure 11.15 DNA Polymerase Binds to the Template Strand (Part 2)
11.3 How Is DNA Replicated?
A primer is required to start DNA
replication—a short single strand of
RNA.
Primer is synthesized by primase.
Then DNA polymerase begins adding
nucleotides to the 3′ end of the primer.
Figure 11.16 No DNA Forms without a Primer
11.3 How Is DNA Replicated?
Cells have several DNA polymerases.
One is for DNA replication; others are
involved in primer removal and DNA
repair.
Other proteins are involved in the
replication process.
Figure 11.17 Many Proteins Collaborate in the Replication Complex
11.3 How Is DNA Replicated?
At the replication fork:
The leading strand is pointing in the
“right” direction for replication.
The lagging strand is in the “wrong”
direction.
Synthesis of the lagging strand occurs in
small, discontinuous stretches—
Okazaki fragments.
Figure 11.18 The Two New Strands Form in Different Ways
11.3 How Is DNA Replicated?
Each Okazaki fragment requires a
primer.
The final phosphodiester linkage
between fragments is catalyzed by DNA
ligase.
Figure 11.19 The Lagging Strand Story (Part 1)
Figure 11.19 The Lagging Strand Story (Part 2)
11.3 How Is DNA Replicated?
DNA polymerases work very fast:
They are processive: catalyze many
polymerizations each time they bind to
DNA
Newly replicated strand is stabilized by a
sliding DNA clamp (a protein)
Figure 11.20 A Sliding DNA Clamp Increases the Efficiency of DNA Polymerization
11.3 How Is DNA Replicated?
The new chromosome has a bit of single
stranded DNA at each end (on the
lagging strand)—this region is cut off.
Eukaryote chromosomes have repetitive
sequences at the ends called
telomeres.
Figure 11.21 Telomeres and Telomerase
11.3 How Is DNA Replicated?
Human chromosome telomeres
(TTAGGG) are repeated about 2500
times.
Chromosomes can lose 50–200 base
pairs with each replication. After 20–30
divisions, the cell dies.
11.3 How Is DNA Replicated?
Some cells—bone marrow stem cells,
gamete-producing cells—have
telomerase that catalyzes the addition
of telomeres.
90% of human cancer cells have
telomerase; normal cells do not. Some
anticancer drugs target telomerase.
11.4 How Are Errors in DNA Repaired?
DNA polymerases make mistakes in
replication, and DNA can be damaged
in living cells.
Repair mechanisms:
• Proofreading
• Mismatch repair
• Excision repair
11.4 How Are Errors in DNA Repaired?
As DNA polymerase adds a nucleotide to
a growing strand, it has a proofreading
function—if bases are paired incorrectly,
the nucleotide is removed.
Figure 11.22 DNA Repair Mechanisms (A)
11.4 How Are Errors in DNA Repaired?
The newly replicated DNA is scanned for
mistakes by other proteins.
Mismatch repair mechanism detects
mismatched bases—the new strand has
not yet been modified (e.g., methylated
in prokaryotes) so it can be recognized.
If mismatch repair fails, the DNA is
altered.
Figure 11.22 DNA Repair Mechanisms (B)
11.4 How Are Errors in DNA Repaired?
DNA can be damaged by radiation, toxic
chemicals, and random spontaneous
chemical reactions.
Excision repair: enzymes constantly
scan DNA for mispaired bases,
chemically modified bases, and extra
bases—unpaired loops.
Figure 11.22 DNA Repair Mechanisms (C)
11.5 What Are Some Applications of Our Knowledge of DNA Structure and
Replication?
Copies of DNA sequences can be made
by the polymerase chain reaction
(PCR) technique.
PCR is a cyclical process:
• DNA fragments are denatured by
heating.
• A primer, plus nucleosides and DNA
polymerase are added.
• New DNA strands are synthesized.
Figure 11.23 The Polymerase Chain Reaction
11.5 What Are Some Applications of Our Knowledge of DNA Structure and
Replication?
PCR results in many copies of the DNA
fragment—referred to as amplifying the
sequence.
Primers are 15–20 bases, made in the
laboratory. The base sequence at the 3′
end of the DNA fragment must be known.
11.5 What Are Some Applications of Our Knowledge of DNA Structure and
Replication?
DNA polymerase that does not denature
at high temperatures (90°C) was taken
from a hot springs bacterium, Thermus
aquaticus.
11.5 What Are Some Applications of Our Knowledge of DNA Structure and
Replication?
DNA sequencing determines the base
sequence of DNA molecules.
Relies on altered nucleosides with
fluorescent tags that emit different
colors of light.
DNA fragments are then denatured and
separated by electrophoresis.
Figure 11.24 Sequencing DNA (Part 1)
Figure 11.24 Sequencing DNA (Part 2)