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Chargaff’s Rules
Erwin Chargaff became interested in
DNA in 1944; he showed that for all
human DNA, the percentages of guanine
and cytosine are nearly identical, 19.1 and
18.4%, respectively, and the percentages
of adenine and thymine, 31.0 and 31.5%,
respectively, likewise are almost identical.
From this remarkable observation, Erwin
Chargaff concluded that the bases always
come in pairs; adenine always associated
with thymine and guanine always
associated with cytosine.
Earlier, scientists had held the view that
DNA was composed of a large number of
repeats of GACT and had explained the
deviation of the percentages from 25% as
an experimental error.
Chargaff’s Rules
In addition to his first rule:
%G = %C and %A = %T
Chargaff also postulated that different species have DNA of different
composition, suggesting DNA as the carrier of genetic information.
Pyrimidines
Purines
The Double Helix of DNA
X-ray diffraction pattern of a hydrated
DNA molecule taken in 1952.
This technique is based on the fact that
the electrons of a molecule diffract Xrays at particular angles; the resulting
pattern (e.g., that above) can be used to
solve the structure of a crystal.
Rosalind Franklin – her X-ray
data led Watson and Crick
(below) to the double helix
12.3
The Double Helix of DNA
Based on Rosalind Franklin’s X-ray
diffraction data, Watson and Crick
proposed a molecular model for
DNA.
This model had a double strand of
repeating nucleotides.
Complementary base pairing (AT,
CG) is held in place by hydrogen
bonds (shown in red).
The nature of the base pairing
required that the two strands be coiled
in the shape of a double helix.
12.3
Structure of DNA
Nucleic acids form a double helix:
DNA is a double helix consisting of two strands of polynucleotides. There
are three types of structural features:
1) phosphoric acid and deoxyribose form
a condensation co-polymer;
2) within the polymer strand the bases are
held together by pi-stacking;
3) two strands, running in opposite
directions, are connected to each other
through highly specific hydrogen-bonds
between their organic bases.
H Bonding
Compounds like alcohols, ethers, and amines, which contain lone
pairs and hydrogens on an electronegative atom (O and N), that is
N–H and O–H functions, can form hydrogen bonds
The O atoms of ketones also can form hydrogen bonds with a
suitable H-donor
Hydrogen Bonds
Structure
Alcohols form hydrogen bonds (cf. H2O)
Alcohols are both H-bond donor and H-bond acceptor
This type of association raises the boiling point
considerably (unbelievably), e.g.,
H3C–CH3 –88°
CH3OH 65°
HDR-F-2012
DNA bases contain lone pairs on N and C=O functions as well as
N–H groups; therefore they can form hydrogen bonds
Pyrimidines
Purines
HDR-F-2012
Place cytosine to the right of guanine and rotate cytosine by -30°
and guanine by 30°
HDR-F-2012
Place thymine to the right of adenine and rotate thymine by -30° and
adenine by 30°
HDR-F-2012
HDR-F-2012
Component Bases of DNA
Structure of DNA
Nucleic acids form base pairs
The individual polynucleotide strands are hydrogenbonded to each other through their organic bases; the
strands run in opposite directions.
Structure of DNA
If a piece of DNA in one strand has the sequence –AGCTACGATC-,
the complementary strand has the sequence
–TCGATGCTAG-
The process by which copies of DNA
are made is called replication.
The original DNA double helix
partially unwinds and the two
complementary portions separate.
Each of the strands serves as a
template for the synthesis of a
complementary strand.
The result is two complete and
identical DNA molecules.
A complete set of genetic
information is packaged into
chromosomes packed into the cell
nucleus.
12.3
Replication of DNA
DNA replicates by unwinding and assembling new
complementary strands.
The double helical nature of DNA suggested to Watson and Crick a mechanism
for the replication of DNA during cell division.
Each of the two strands in a
parent DNA molecule serve
as a template for the assembly
of a new complementary
strand in a new DNA
molecule. This process occurs
with an error frequency of
less than 1 in 10 billion base
pairs during DNA replication.
The nearly 3 billion base pairs in each human cell provide the
blueprint for producing a human being.
The specific sequence of base pairing is important in
conveying the mechanism of how genetic information is
expressed.
The expression is enacted through proteins.
DNA directs the synthesis of proteins and, thereby, controls the
characteristics of an individual, including inherited diseases.
Let’s review Proteins
12.4
Proteins
Proteins are made of amino acids. The general formula for an amino acid includes
four groups attached to a carbon atom: (1) a carboxylic acid group, –COOH; (2) an
amine group, –NH2; (3) a hydrogen atom, –H; and (4) a side chain
There are 20 naturally
occurring amino acids
that make up proteins.
They differ from one another by the different R groups
12.4
Proteins
Two amino acids can link together via a peptide bond:
amino acid residue
The two molecules join,
expelling a molecule of water.
Peptide bond
As the process repeats itself again and again, it creates a
peptide chain.
Once incorporated into the peptide chain, the amino acids
are known as amino acid residues.
12.4
The Genetic Code
The order of amino acids in a protein, the genetic code, is determined by the order
of bases in DNA (Marshal Nirenberg, Nobel Prize, Phys. & Med. 1968).
Because human protein contains 20 amino acids, the DNA code must contain 20
code “words”, each representing a specific amino acid. The four DNA bases can be
combined to a sufficient number of three DNA base groups, called codons, to
represent each amino group uniquely; this sequence of codons is the genetic code.
The diagram shows three codons, GTA, AAA, and GGC, which for the
incorporation of histidine, phenylalanine, and proline, respectively. Some amino
acid residues can be produced by different codes
AAA
lysine
CCC
proline
GGG
glycine
Protein Structure
The primary structure of a protein is its linear sequence of amino
acids, including the location of any disulfide (–S–S–) bridges.
12.5
Protein Structure
The secondary structure of a protein is the folding pattern
within a segment of the protein chain.
12.5
Protein Structure
N-terminal
carboxyl
terminal
The sequence is
characterized by the
amino terminal or
"N-terminal" (NH3+) at
one end, and the
carboxyl terminal or
"C-terminal" (COO–) at
the other.
Tertiary structure of the enzyme, chymotrypsin
12.5
Protein Function and Structure
The function of a protein is dependent on its shape or threedimensional structure. Small changes in the primary structure can
have dramatic effects on its properties.
Sickle cell anemia is an example of
a condition that develops when red
blood cells take on distorted
shapes due to an error in the amino
acid sequence.
Because of their irregular shape
these cells cannot pass through
tiny openings in the spleen and
other organs.
Some of the sickled cells are destroyed and anemia results. Other
sickled cells can clog organs so badly that the blood supply to them
is reduced.
12.5
Genetic Engineering
•  When a species is genetically engineered, the DNA in the cell is modified.
•  When the genes are changed, the proteins synthesized by the genes are
modified.
•  When the cell grows and develops, a plant with new characteristics from the
different DNA is generated.
•  Before genetic engineering, when humans
selected for plants with certain characteristics
or crossbred different strains, genes were
manipulated.
The genetic traits for modern corn were selected over time, starting with an early
ancestor, teosinte (bottom).
12.6
Genetic Engineering
12.6
Genetic Engineering
•  Genetic engineering is transgenic – where an organism is created
by the transfer of genes across species.
•  Genetic engineering can also be used to do the same thing as
crossbreeding, just more efficiently and faster.
Transgenic rice with virus-resistance
12.6
Reasons for Genetic Engineering
•  Make crops more resistant to disease, tolerant of stresses (salt, heat,
or drought)
•  Develop soybeans that produce high yields of biofuel per acre
•  Use of enzymes to create new drugs
•  Develop vaccines that grow in edible products
Developing strains of algae for new biofuels
12.7
Genetically-Engineered Agriculture
Transgenic Plants
Virus resistant transgenic rice
Frankenfood?
Greenpeace activists dumping
papaya during a Bangkok protest.
12.8
Genetic Engineering and Sight
Recombinant DNA used to restore sight to
children with congenital blindness.
A research team at the Univ. of Pennsylvania
School of Medicine created a vector (a genetically
engineered virus) to carry a normal version of a
gene called RPE65, that is mutated in one form of
Leber’s congenital amaurosis (LCA), a genetic
disease that progressively damages the retinas
leaving many patients totally blind in their twenties
or thirties.
Genetic Engineering
Animal studies with mice and dogs had shown
that visual improvement was age-dependent, so the
research team hypothesized that younger patients
would receive the greatest benefit. A clinical trial
with five children and seven adults ranging in age
from 8 to 44, received injections of therapeutic
genes into their retinas.
Genetic Engineering
As expected, the greatest improvement
occurred in the children, all of whom are now
able to navigate a low-light obstacle course.
Before they received the gene therapy, the
patients had great difficulty avoiding barriers,
especially in dim light.
Not all the adults performed better on the
obstacle course and those who did, showed
more modest improvements than did the
children.
Genetic Engineering
The clinical benefits have persisted for
nearly two years after the first injections with
therapeutic genes were given. Although none
of the patients attained normal eyesight, six of
the twelve test subjects improved enough that
they are no longer classified as legally blind.
Gene therapies for other retinal diseases,
such as age-related macular degeneration may
also be developed.
Cloning Mammals and Humans
In 1996, Dolly the sheep was born – the first
cloned mammal. Dolly was created by a technique
called nuclear transfer. The nucleus (containing the
chromosomes) from an adult cell was placed in a
donor egg from another sheep whose nucleus had
been removed. The nucleus and donor egg were
fused with an electrical jolt. The DNA then initiated
the growth of the embryo, which was then
implanted into a surrogate sheep’s uterus.
Since then, several other mammalian species
have been successfully cloned.
Cloning Mammals and Humans
Dolly is an example of “reproductive
cloning”, in which an embryo is transferred to a
gestational carrier in the hopes that a
pregnancy will result and be carried full term.
Cloning
Mammals
and
Humans
Cloning Humans and Mammals
Cloning Mammals and Humans
Cloning Mammals and Humans
Cloning Humans and Mammals
Cloning Humans and Mammals
Dolly, the cloned sheep
Dolly, the cloned sheep
Snuppy, the cloned dog, next to his “father
Dolly, a cloned sheep,
Snuppy,
a next
cloned
dog,
Snuppy, the
cloned dog,
to his “father”
next to his “father”
Therapeutic Cloning
“Therapeutic cloning” refers to harvesting
stem cells from 3- to 5-day-old embryos to establish
stem cell lines. Scientists hope to induce these stem
cells to differentiate into various specialized cells.
In 2004, a team of scientists led by Woo Suk
Hwang and Shin Yong Moon of Seoul National
University reported that they had successfully
cloned human cells to generate embryonic stem
cells. In 2006, Hwang admitted the data had been
fabricated and resigned from his university position.
Therapeutic Cloning
Would blastocysts created in this manner be
extensions of the people whose DNA was used to
create them or would they be separate, unique
beings in the same way that identical twins are
unique, even though they share the same genetic
blueprint?
Stem Cell Research
Scientists at the Burnham Institute for
Medical Research in La Jolla, CA, have
programmed embryonic stem cells into becoming
nerve cells when transplanted into the brains of
mice. None of the mice formed tumors, which have
been a major setback in previous attempts at stem
cell transplantation.
This research is a first step toward developing
new treatments for stroke, Alzheimer’s, and other
neurological conditions.
Stem Cell Research
The Food & Drug Administration (FDA)
approved a phase I clinical trial for the
transplantation of a human embryonic stem cellderived cell population into spinal cord-injured
individuals on January 23, 2009.
Eight to ten paraplegics who were injured less
than two weeks before the trial begins, will be
selected, because the neural stem cells must be
injected before scar tissue forms. These first trials
are mainly to test for the safety of the procedures.
Stem Cell Research
Based on earlier results with mice, researchers
say the restoration of myelin sheaths (insulation
around nerve cells) and an increase in mobility is
probable. The injections are not expected to fully
restore mobility.
In November 2010, the first patient, a recent
paraplegic, was injected with two million
embryonic stem cells in the injured spinal cord
region with the goal of regenerating spinal cord
tissue. The cells had been induced to become
specialized nerve cells.
Stem Cell Research
The embryonic stem cells came from a
leftover embryo from a fertility treatment, which
would have been otherwise discarded. Embryonic
stem cells are valued by researchers for their ability
to be transformed into any type of cell. There are
some restrictions tied to federally funded research
involving embryonic stem cell lines. The company
developing this treatment has spent ~$175 million
thus far without federal funding.
Animal model studies have shown movement
in previously paralyzed rodents, but the results in
humans are not expected to be that dramatic.
Stem Cell Research
Human embryonic stem cells could be used as
models for human genetic diseases. The relative
inaccessibility of human tissue is an obstacle to
research in these areas. This approach could be very
valuable in studying cystic fibrosis or fragile-X
syndrome or other genetic diseases where no
reliable animal model exists.
Embryos with a genetic disease could be
identified by prenatal genetic diagnosis (PGD) and
used to establish a stem cell line featuring the
genetic disorder.
Stem Cell Research
Human embryonic stem
cells are grown typically
on a mouse feeder layer;
they thrive on it
Stem Cell Research
Stem Cell Research
The Busch administration did not allow the
National Institutes of Health (NIH) to provide any
funding for introduction of new human embryonic
stem cell lines.
On Dec. 2, 2009, the NIH announced the
approval of thirteen new human embryonic stem
cell lines for NIH funding, still a strictly limited
number.
Genetic Engineering
Where do we go from here?
Is saving a human life worth the
cost of a potential human life?
12.8