Download Ch9_DNA

DNA
The Universal Code of Life
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• To which class of biological molecules does
DNA belong?
• What are the monomers of DNA?
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DNA is a:
90%
1.
2.
3.
4.
5.
Carbohydrate
Protein
Lipid
Nucleic Acid
Depends on the
organism
9%
2%
1
0%
2
3
0%
4
5
The monomers of DNA are:
98%
1.
2.
3.
4.
5.
Sugars
Amino acids
Fatty acids
Nucleotides
Depends on the
organism
2%
1
0%
0%
2
3
0%
4
5
Early History
• 1869: Friederich Mieschner isolates
“nuclein” from nuclei of cells. His student
Richard Altman later renames the substance
“nucleic acid.”
• Mid 1800s: Biochemists identify two distinct
nucleic acids.
• 1929: Phoebus Levine identifies four distinct
bases in DNA.
Heredity as a Science
• Genetics arose as a new science in the late
19th and early 20th centuries, spurred by
questions raised by Darwin’s On the Origin
of Species:
• Are there patterns to inheritance?
• Are traits handed on intact (particle
theory) or blended together in each
generation (blending theory)?
Mendel’s Answers
• Gregor Mendel’s work was rediscovered in
1900, answering both questions:
• Inheritance of many traits follows
predictable patterns.
• Traits are handed on intact via some kind
of particle: “elementen.”
Hereditary Molecule?
• Question in the 20th century: What is the
hereditary molecule?
• Cell nucleus associated with inheritance.
• Both proteins and nucleic acids are in the
nucleus. Which contains information
coding for traits?
Protein or DNA?
• Linus Pauling favored protein: DNA has
only four bases, protein has over 20 amino
acids. Seemed like protein could store more
information.
• Others favored DNA, which is found only in
the nucleus.
Frederick Griffith
• In 1928, Frederick Griffith
carried out experiments on
pneumonia bacteria, trying to
create a vaccine against
pneumonia. Among his findings
were early clues about hereditary
factors.
Griffith’s Experiment
Bacterial strain(s) injected into mouse
Results
Mouse remains
healthy.
Living
R-strain
Conclusions
R-strain does not
not cause
pneumonia.
Mouse contracts
pneumonia, dies.
Living
S-strain
Heat-killed
S-strain
Living R
strain,
heat-killed
S-strain
S-strain causes
pneumonia.
Mouse remains
healthy.
Heat-killed Sstrain does not
cause pneumonia.
Mouse contracts A substance from
pneumonia, dies. heat-killed S-strain
can transform the
harmless R-strain
into a deadly
S-strain.
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Why were living S-strain
bacteria recovered from dead
mice injected with dead Sstrain and live R-strain
bacteria?
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Oswald Avery
• Avery learned of Griffith’s experiment and
thought it might hold a clue to the identity of
the hereditary molecule.
• Avery isolated carbohydrates, proteins,
lipids, and nucleic acids from the bacteria to
discover which, if any, would transform the
non-virulent R-strain bacteria.
Of the substances isolated and tested, only DNA from
killed S-strain bacteria transformed R-strain bacteria.
Hershey & Chase
• Early 1950’s: Alfred Hershey
and Martha Chase used the
bacteriophage virus in another
series of experiments to
identify the hereditary
material.
• Bacteriophages, like other
viruses, contain both protein
and DNA, but are non-living.
DNA
head
Bacteriophage
Protein coat
tail
6 Bacterial wall
destroyed; phage
released.
5 Complete
phages
assembled.
4 Phage parts
synthesized, using
bacterial metabolism.
1 Phage attaches
to bacterium.
2 Phage
injects its
DNA into
bacterium.
3 Phage DNA
is replicated.
Radio-tagged DNA
Radio-tagged Protein
Radioactive phosphorus (P32)
Radioactive sulfur (S35)
Radioactive
DNA (blue)
1 Label phages with P32 or S35.
Radioactive
protein
(yellow)
2 Infect bacteria with
labeled phages; phages inject
genetic material into bacteria.
3 Whirl in blender to break off
phage coats from bacteria.
4 Centrifuge to separate phage coats
(low density: stay in liquid)
from bacteria (high density:
sink to bottom as a “pellet”)
5 Measure radioactivity of phage
Results: Bacteria are
Results: Phage coats are
coats and bacteria.
radioactive; phage coats are not.
radioactive; bacteria are not.
Conclusion: Infected bacteria are labeled with radioactive phosphorus but not with radioactive sulfur,
supporting the hypothesis that the genetic material of bacteriophages is DNA, not protein.
These early experiments showed that DNA
is the hereditary molecule because:
1. Only DNA could
break down
proteins.
2. Only DNA caused
changes in
hereditary traits.
3. Only bacteria and
viruses have DNA.
93%
7%
1
0%
2
3
DNA Structure?
• While many research teams were trying to
discover the hereditary molecule, other
researchers were working to discover the
nature of DNA.
Erwin Chargaff
• Chargaff took apart DNA into
its component nucleotides and
studied the proportions.
• Found consistent ratios
between certain nucleotides.
In DNA, Chargaff
consistently found
equal amounts of
adenine compared with
thymine, and equal
amounts of cytosine
compared with guanine.
Did that mean the bases
were always paired?
If a strand of DNA is 30% adenine, how
much thymine does it have?
95%
1.
2.
3.
4.
15%
20%
30%
Impossible to
predict
2%
1
2%
2
2%
3
4
If a strand of DNA is 30% adenine,
how much cytosine does it have?
62%
1.
2.
3.
4.
15%
20%
30%
Impossible to
predict
12%
9%
1
2
3
17%
4
Franklin and Wilkins
• Rosalind Franklin worked in Maurice
Wilkins’ lab in the late 1940s, using X-ray
crystalography to find clues about the
structure of DNA.
• Franklin’s images were the first to suggest a
helical structure.
The X-shape on the radiograph was characteristic of helical
molecules. Franklin also measured distances between
bases and other dimensions using her images.
Watson and Crick
• James Watson and Francis Crick worked at
the same time as Franklin and Wilkins.
• Applying Chargaff’s rule, they concluded
that A pairs with T, C with G.
• Used their knowledge of molecular
geometry to try to discover the structure of
DNA.
• Wilkins consulted with
Watson and Crick.
Without Franklin’s
knowledge, he handed
them several of
Franklin’s X-ray images.
• Watson immediately
recognized their
significance, though he’d
criticized Franklin’s
work earlier.
By adding Franklin’s data to their own (without her
permission!), Watson and Crick assembled the first
plausible model of DNA and published an article on the
structure of DNA in 1953.
DNA Structure
DNA contains four bases. RNA also has four bases, but has
uracil instead of thymine.
How many
rings?
How many
rings?
How many
H-bonds?
How many
H-bonds?
As Chargaff’s work suggested, Adenine always pairs across
the DNA ladder with Thymine, while Cytosine always pairs
with Guanine.
5’ end
5 4
6
5’
1’
4’
3’
3
1 2
2’
3’ end
Nucleotides are 3-dimensional, with an orientation that
affects the shape of the entire nucleic acid.
5’ end
5’
1’
4’
3’
5’
3’
3’ end
5 4
6 3
1 2
2’
1’
4’
The 3’ end of one
nucleotide binds with
the 5’ end of the next
nucleotide in the chain.
2’
8
9
7
5 4
6 3
1 2
5’end
3’ end
Two chains of DNA
nucleotides are held
together by
hydrogen bonds
between the bases
of each strand.
Notice that the strands
run in opposite
directions. They are
antiparallel.
3’end
5’end
free phosphate
free sugar
The 3-dimensional shape of
the nucleotides creates the
helical structure of DNA.
The sugar in the backbone of
DNA is:
62%
1.
2.
3.
4.
Glucose
Ribose
Deoxyribose
Lactose
16%
19%
3%
1
2
3
4
In the DNA double helix, adenine always
matches thymine because:
89%
1. Adenine is polar and
thymine is nonpolar.
2. Both can form two
hydrogen bonds with
each other.
3. Both are single-ring
bases.
4. Wrong! Adenine always
matches adenine.
11%
0%
1
0%
2
3
4
• Suppose that one side of a DNA doublehelix reads:
ATAACAGTTAGCAGG
According to the base-pairing rule, what is
the sequence of bases on the other side of the
DNA double-helix?
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Label the four bases in
this diagram. (Look
back several slides for
a hint.)
A
A
G
C
T
C
G
Circle one complete
nucleotide on each
side. (Hint: look back
several slides to see
which carbon on the
sugar attaches to the
phosphate.)
DNA Replication
• When cells divide, the two resulting
daughter cells must have exactly the same
DNA as the original cell.
• Therefore, before cell division happens, the
cell must replicate (copy) its DNA.
replication bubbles
DNA
DNA helicase
replication forks
The enzyme DNA helicase “unzips” DNA by breaking
hydrogen bonds holding the two strands together.
“Unzipping” occurs at multiple points on the DNA strand.
DNA helicase
replication forks
DNA
polymerase #1
3
5
3
DNA
polymerase #2
5
Within each replication bubble, the enzyme DNA
polymerase builds a new strand of DNA, using the original
strands as templates.
DNA
polymerase #1
3
5
3
DNA
polymerase #2
DNA polymerase #1
continues along
parental DNA strand
5
3
DNA
polymerase #3
5
3
DNA
5 polymerase #2
leaves
3
5
Because DNA polymerase always travels from the 3’ to the
5’ end of DNA, one polymerase is always moving away from
the replication fork
DNA polymerase #1
continues along
parental DNA strand
5
3
DNA
polymerase #3
5
3
DNA
polymerase #4
3
DNA
5 polymerase #2
leaves
3
5
3
5
3
DNA
polymerase #3
leaves
5
3
5
DNA ligase joins
daughter DNA strands
together.
Multiple DNA polymerase molecules are required for the
strand where discontinuous replication is happening.
How does DNA polymerase “know” which bases to use
when replicating?
Remember Chargaff’s
rule: A and T always
match, C and G always
match.
Practice DNA base-pair matching:
http://learn.genetics.utah.edu/content/begin/dna/builddna
• Suppose a segment of a DNA double-helix
reads:
AGTCAATGC
TCAGTTACG
After replication, what will the two resulting
DNA double-helices read?
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The enzyme that “unzips” DNA is:
33%
33%
33%
1. DNA polymerase
2. Helicase
3. Ligase
1
2
3
The enzyme that “pastes” in new
bases during replication is:
33%
33%
33%
1. DNA polymerase
2. Helicase
3. Ligase
1
2
3
The enzyme that mends gaps in the
sugar-phosphate backbone is:
33%
33%
33%
1. DNA polymerase
2. Helicase
3. Ligase
1
2
3
• Helicase, DNA polymerase, and ligase are
enzymes. To which class of biological
molecules do enzymes belong?
• Where are the instructions for making DNA
polymerase found?
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Mutations
• Though many enzymes patrol your DNA,
looking for replication errors, some errors do
creep in.
• Most cells with a DNA error will die. A few
may turn cancerous.
• If mutated cells are sex cells, the mutation
can be passed on and will affect all cells in
the offspring.
• Mutations may be harmful, helpful, or
neutral.
• Harmful mutations result in genetic
disease or death.
• Helpful mutations increase evolutionary
“fitness” (i.e. having offspring).
• Neutral mutations neither help nor harm at
the present.
Nucleotide substitution
original DNA sequence
substitution
nucleotide pair changed
from A–T to T–A
Silent mutation: still
codes for the same
amino acid.
Missense mutation: codes
for a different amino acid,
which may or may not
affect the final protein.
Nonsense mutation: codes for
a “stop” in the middle of the
sequence, producing a
useless protein.
Examples:
• Sickle-cell anemia is caused by a missense
mutation due to a nucleotide substitution.
• Duschenne’s Muscular Dystrophy is caused by
a nonsense mutation in a gene for a critical
enzyme.
• Lactose persistence may be caused by a single
nucleotide substitution.
Insertion mutation
original DNA sequence
Example: Huntington’s
disease, a loss of neural
function in middle-age, is
caused by a string of
insertions.
T–A nucleotide pair
inserted
Deletion mutation
original DNA sequence
Example: a 32-base-pair
deletion in the gene for a certain
cell membrane receptor protein
causes resistance to HIV.
C–G nucleotide pair
deleted
Inversion
original DNA sequences
breaks
DNA segment inverted
Example: an inversion
mutation is responsible for
one form of hemophilia.
Translocation
original DNA sequences
break
DNA
segments
Switched
break
Examples: Several forms of leukemia, lymphoma
and possibly schizophrenia are caused by
translocation mutations.
What causes mutations?
http://learn.genetics.utah.edu/archive/sloozeworm/mutation
bg.html
Recap
• DNA is a nucleic acid which contains coded
hereditary information.
• The base-pairing rule helps the information
in DNA accurate.
• All cells in the body have the same DNA
containing the same information. DNA must
be replicated before cell division.