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DNA (Deoxyribonucleic Acid)
Scientific History

The march to understanding that DNA is
the genetic material
– T.H. Morgan (1908)
– Frederick Griffith (1928)
– Avery, McCarty & MacLeod (1944)
– Erwin Chargaff (1947)
– Hershey & Chase (1952)
– Watson & Crick (1953)
– Meselson & Stahl (1958)
Transformation of Bacteria
What carries hereditary information?





By the 1940s, scientists knew that
chromosomes carried genes.
They also knew that chromosomes were
made of DNA and protein.
They did NOT know which of these
molecules actually carried the genes.
Since protein has 20 types of amino acids
that make it up, and DNA only has 4 types
of building blocks, it was a logical
conclusion.
Most Scientists thought protein carried
genes
Chromosomes are made of DNA and protein
Transformation of Bacteria
DNA is the “Transforming
Principle”

1944
Avery, McCarty & MacLeod
– purified both DNA & proteins separately from
Streptococcus pneumonia bacteria
• which will transform non-pathogenic bacteria?
– injected protein into bacteria
• no effect
– injected DNA into bacteria
• transformed harmless bacteria into
virulent bacteria
mice die
What’s the
conclusion?
Avery’s Experiment
1. Avery repeated Griffith’s
experiments with an
additional step to see what
type of molecule caused
transformation.
3. When Avery added
enzymes that destroy
DNA, no transformation
occurred.
So…he knew that
DNA carried
hereditary
information!
2. Avery used enzymes to destroy the sugars and
transformation still occurred—Sugar did not cause
transformation.
Avery used enzymes to destroy lipids, RNA, and
protein one by one. Every time transformation still
occurred—none of these had anything to do with
the transformation.
1944 | ??!!
Avery, McCarty & MacLeod

Conclusion
– first experimental evidence that DNA was the genetic
material
Oswald Avery
Maclyn McCarty
Colin MacLeod
Hershey-Chase Experiment




The experiment involved viruses
to see if DNA or protein was
injected into the bacteria in
order to make new viruses.
One group of viruses was
infected with radioactive
protein and another group with
radioactive DNA.
Then the viruses attack the
bacteria.
Radioactive DNA shows up in
the bacteria, but no
radioactive protein.
1947
Chargaff

DNA composition: “Chargaff’s rules”
– varies from species to species
– all 4 bases not in equal quantity
– bases present in characteristic ratio
• humans:
A = 30.9%
T = 29.4%
G = 19.9%
C = 19.8%
interesting!
That’s
What do you notice?
Rules
A = T
C = G
Rosalind Franklin


Took X-ray
pictures of DNA.
The photos
revealed the
basic helix,
spiral shape of
DNA.
Maurice Wilkins


Worked with
Rosalind Franklin.
Took her x-ray
photos and
information to
Watson and Crick
Watson and Crick



Used Franklin’s pictures
to build a series of
large models.
Stated that DNA is a double-stranded molecule in
the shape of a double helix, or twisted ladder.
Won the Nobel Prize for their work in 1962.
Semiconservative replication,


when a double helix replicates each of the daughter molecules will
have one old strand and one newly made strand.
Experiments in the late 1950s by Matthew Meselson and Franklin
Stahl supported the semiconservative model, proposed by Watson
and Crick, over the other two models. (Conservative & dispersive)
Double helix structure of DNA
“It has not escaped our notice that the specific pairing we have postulated
immediately suggests a possible copying mechanism for the genetic
material.”
Watson & Crick
Basic DNA Structure
P

S
A
P

S
C
P
S
T
A nucleotide is
the monomer of
DNA
A nucleotide is
made of
– a sugar called
deoxyribose
– a phosphate
– and a base
(ATCG)
Directionality of DNA

You need to
number the
carbons!
nucleotide
PO4
N base
– it matters!
5 CH2
This will be
IMPORTANT!!
O
4
1
ribose
3
OH
2
Deoxyribose


Simple sugar
molecule like
glucose that has 5
carbons
The five carbons are
numbered clockwise
starting from the first
one after the oxygen
Phosphate

The negatively charged phosphate
bonds to the 5’ Carbon of the
deoxyribose.
Bases

The base bonds to the 1’ Carbon.
Base
Bases

There are two main types of bases
purines and pyrimidines.
– Purines have two rings in their structure.
• Adenine and guanine are purines.
– Pyrimidines only have one ring.
• Thymine and Cytosine are pyrimidines.
Pyrimidines
Purines
Basic DNA Structure
P

S
A
S
C
S
T
P
P
To form one strand
of DNA, the
phosphate of one
nucleotide covalently
bonds to the 3’
Carbon of the
deoxyribose from
another nucleotide.
P
S
A
T
S
P
P
S
C
G
S
P
P
S
T
A
S
P

The two strands of DNA are held
together by hydrogen bonds
Anti-parallel strands

Nucleotides in DNA
backbone are bonded from
phosphate to sugar between
3 & 5 carbons
5
3
3
5
– DNA molecule has “direction”
– complementary strand runs in
opposite direction
Base Pairs

The nucleotides that bond together by
their bases are called base pairs.
– Adenine only bonds to Thymine
– Guanine only bonds to Cytosine
Does each of your cells
have the same DNA?
YES
DNA Replication

Before a cell divides,
DNA must make a copy
of itself so that each
new cell has a
complete set of DNA.
Step 1-Unzip DNA

An enzyme called helicase untwists the
ladder and breaks the hydrogen bonds
between the bases and “unzips” DNA
down the middle.
Helicase Enzyme
Step 2-Prime the DNA


An enzyme called DNA primase put a
few nucleotides of RNA on the DNA.
This is only to create a starting place
and these will later be removed.
Step 3-Elongation


The two strands of the
Parent DNA become
templates for the new
strands.
New nucleotides are
added by an enzyme
called DNA polymerase.
Step 3-Elongation

DNA polymerase
only adds
nucleotides in the 5’
to 3’ direction on
both strands
beginning at the
RNA primer.
Step 4 – Fine tuning


RNA primer is removed and any gaps
are sealed by an enzyme called ligase.
DNA polymerase proof reads the new
copy and fixes any mistakes.
Helicase unwinds and unzips DNA
P
S
A
T
S
P
P
S
C
G
S
P
P
S
T
A
S
P
DNA Polymerase Adds New Nucleotides
P
P
S
A
T
S
S
P
P
S
C
G
S
T
A
P
C
G
S
P
P
S
S
S
P
S
P
T
P
S
P
A
T
A
S
P

Are the two copies of DNA the
same?

Why would it be important for the two
copies of DNA to be the same?
Okazaki
Leading & Lagging strands
Limits of DNA polymerase III

can only build onto 3 end of an
existing DNA strand
5
3
5
3
5
3
5
5
5

Lagging strand
ligase
growing
3
replication fork
Leading strand

3
Lagging strand


3
Okazaki fragments
joined by ligase
 “spot welder” enzyme
5
3
DNA polymerase III
Leading strand

continuous synthesis
Replication fork / Replication bubble
3
5
5
3
DNA polymerase III
leading strand
5
3
3
5
3
5
5
5
3
lagging strand
3
5
3
5
lagging strand
5
5
leading strand
growing
replication fork 5
3
growing
replication fork
leading strand
3
lagging strand
5 5
5
5
3
Starting DNA synthesis: RNA
primers
Limits of DNA polymerase III

can only build onto 3 end of an
existing DNA strand
5
3
3
5
5
3
5
3
5
growing
3
replication fork
DNA polymerase III
primase
RNA 5
RNA primer


built by primase
serves as starter sequence for DNA
polymerase III
3
Starting DNA synthesis: RNA
primers
Limits of DNA polymerase III

can only build onto 3 end of an
existing DNA strand
5
3
3
5
5
3
5
3
5
growing
3
replication fork
DNA polymerase III
primase
RNA 5
RNA primer


built by primase
serves as starter sequence for DNA
polymerase III
3
Replacing RNA primers with DNA
DNA polymerase I

removes sections of RNA primer and
DNA polymerase I
replaces with DNA nucleotides
5
3
3
5
5
ligase
growing
3
replication fork
RNA
5
3
But DNA polymerase I still
can only build onto 3 end of
an existing DNA strand
Houston, we
have a problem!
Chromosome erosion
All DNA polymerases can
only add to 3 end of an
existing DNA strand
DNA polymerase I
5
3
3
5
5
growing
3
replication fork
DNA polymerase III
RNA
Loss of bases at 5 ends
in every replication


chromosomes get shorter with each replication
limit to number of cell divisions?
5
3
Telomeres
Repeating, non-coding sequences at the end
of chromosomes = protective cap

limit to ~50 cell divisions
5
3
3
5
5
growing
3
replication fork
telomerase
5
Telomerase



enzyme extends telomeres
can add DNA bases at 5 end
different level of activity in different cells
 high in stem cells & cancers -- Why?
TTAAGGG TTAAGGG 3
Replication fork
DNA
polymerase III
lagging strand
DNA
polymerase I
5’
3’
ligase
primase
Okazaki
fragments
5’
3’
5’
SSB
3’
helicase
DNA
polymerase III
5’
3’
leading strand
direction of replication
SSB = single-stranded binding proteins
Length of DNA

The length of the DNA from one cell is
– 3 meters

"Unravel your DNA and it would stretch
from here to the moon"
DNA Packing
DNA
double
helix
(2-nm
diameter
Histones
“Beads on
a string”
Nucleosome
(10-nm diameter)
Tight helical fiber
(30-nm diameter)
Supercoil
(200-nm diameter)
700
nm
Metaphase chromosome
Nucleosomes

“Beads on a string”
– 1st level of DNA packing
– histone proteins
• 8 protein molecules
• positively charged amino acids
• bind tightly to negatively charged DNA
8 histone
molecules
DNA packing as gene control

Degree of packing of DNA regulates transcription
– tightly wrapped around histones
• no transcription
• genes turned off
 heterochromatin
darker DNA (H) = tightly packed
 euchromatin
lighter DNA (E) = loosely packed
H
E
The End!