Download DNA I. History of the Role of DNA Since the late 1800`s, scientists

I.
DNA
History of the Role of DNA
Since the late 1800’s, scientists knew that information was
passed from parent to offspring in a predictable way. As
biology and genetics advanced, scientists were able to begin
narrowing down exactly which substance in the cell was
responsible for containing the genetic information. From
1910’s to the 1940’s, there was a debate between whether
nucleic acids or proteins were the source of genetic
information. And then a few key experiments in the first half
of the 1900’s shed some light on the subject.
A. 1928: Frederick Griffith – Bacterial Transformation
1. Griffith was trying to figure out how bacteria make
people sick and was studying pneumonia.
a. Had isolated two different strains of bacteria
(same species, different variety) in mice
b. S strain – caused pneumonia in mice, colonies
have smooth edges
c. R strain – did not cause disease in mice, colonies
have rough edges
2. Griffith’s Experiments
a. The following experiments were done as
controls:
i. Living S bacteria injected into mouse:
mouse develops pneumonia and dies
ii. Living R bacteria injected into mouse:
Mouse stays healthy
iii.
Heat killed S bacteria injected into
mouse:
Mouse stays healthy
b. But when Griffith injected heat-killed S bacteria
mixed with live R bacteria into a mouse, the
mouse developed pneumonia and died! By
themselves, neither type of bacteria should have
made the mouse sick.
i. When the dead mice were examined,
Griffith found that the lungs were full
of S bacteria.
ii. How was this possible if the S bacteria
were dead when he injected them into
the mouse?
Griffith’s Experiments - Transformation
Hereditary material from the
heat-killed S cells transforms
R cells. The transformed R cells
kill the mouse.
c. Somehow, the heat killed S bacteria passed their
disease causing ability to the harmless R
bacteria. Griffith concluded that some chemical
factor transferred from the heat killed S bacteria
to the live R bacteria. He reasoned that the
chemical factor must contain information that
could change the harmless bacteria into the
disease causing bacteria.
d. Griffith also found that the newly disease
causing bacteria passed down the characteristic
to their descendents, so he figured the chemical
factor must be a gene.
e. Griffith called this phenomenon transformation.
f. Transformation: a change in the genes or traits
of an organism due to the addition of outside
DNA by a cell
B. 1944: Oswald Avery – Cause of Transformation
1. Avery and his team wanted to determine which
molecule in the heat killed bacteria was most
important for transformation to occur. If they were
successful, they might help to reveal the chemical
nature of the gene.
2. The experiments:
a. Avery extracted a mixture of various molecules
from the heat killed bacteria, most notably
protein, lipids, carbohydrates, RNA, and DNA.
b. Enzymes were used to separately destroy each of
the molecules in heat killed bacteria. So one
batch of heat killed bacteria had proteins
destroyed, another had the RNA destroyed, and
so on.
c. It was found that after proteins, lipids,
carbohydrates, and RNA were destroyed,
transformation still occurred. It was only after
the DNA was destroyed that transformation did
not happen. Avery concluded that DNA must be
the molecule responsible for transformation.
3. It would take further experimentation for the
scientific community to accept that DNA was
definitely the material of the gene. Some scientists
still felt that protein was more suited to the function
of the gene. The work of Griffith and Avery are the
foundation on which further work would build to
show that the gene was indeed composed of DNA.
C. 1952: Alfred Hershey and Martha Chase – Protein or DNA?
1. Hershey and Chase studied a bacteriophage to find
out if it was indeed DNA or protein that was the
material of the gene.
2. Bacteriophage: a virus that infects bacteria
a. At the time, biologists knew that most viruses were
composed almost entirely of DNA and protein
b. They also knew that the virus would quickly turn
the bacterial cell into a factory for making more
viruses.
c. But they didn’t know how the virus reprogrammed
the cell to produce viruses. Was it the DNA or the
protein that was responsible? Which one actually
entered the bacteria?
3. The Experiment
a. Viruses were grown in cultures containing
radioactive isotopes. Since they are radioactive,
the isotopes could be easily detected and would act
as “tags” for the protein and for the DNA.
b. One culture was grown in radioactive phosphorus
and another in radioactive sulfur. Why these two
elements? Proteins contain almost no phosphorus
and DNA contains no sulfur.
c. So the virus’s protein coat was marked with
radioactive sulfur and the virus DNA was marked
with radioactive phosphorus.
d. The marked viruses were then mixed with bacterial
cells, given time for the viruses to infect the
bacteria, and then separated the viruses from the
bacteria.
e. Hershey and Chase then checked the bacteria for
radioactivity. They found radioactive phosphorus,
the marker on the viral DNA. It was the DNA that
was injected into the bacteria, not the protein.
DNA was the genetic material, not protein.
Alfred Hershey and Martha Chase used different radioactive markers to label the DNA
and proteins of bacteriophages. The bacteriophages injected only the DNA into the
bacteria, not proteins. From these results, Hershey and Chase concluded that the genetic
material of the bacteriophage was DNA.
4. Hershey and Chase confirmed Avery’s results. Their
work was crucial in convincing the scientific
community that DNA was the genetic material found
in cells.
II. Structure of DNA
While Hershey and Chase worked to confirm Avery’s results,
the race was on to find the structure of the gene, or DNA.
We’ll be going into more detail of the key people involved later,
but we’re going to look into the actual structure now.
Structure of DNA
Remember: genes control certain traits, genes are sections of DNA
I. Structure of DNA (deoxyribonucleic acid)
A. Made of nucleotides
1. nucleotides have 3 main parts
a. sugar (deoxyribose)
S
b. phosphate group
P
c. nitrogenous base (see below)
2. 4 different nitrogenous bases can be used in a nucleotide
a. adenine (A)
b. guanine (G)
c. cytosine (C)
d. thymine (T)
3. DNA formed by 2 strands of nucleotides linked together
a. “double helix shape”  twisted ladder
b. sides of the ladder are alternating molecules of sugar
(deoxyribose) and phosphate
c. “rungs” of ladder are 2 bases bonded together by
hydrogen bonds:
i. Adenine always bonds with Thymine – held
by 2 hydrogen bonds
ii. Cytosine always bonds with Guanine – held
by 3 hydrogen bonds
iii. A = T and G = C is known as Chargaff’s
Rule(s) – based on his experiments in 1950
showing that in samples of DNA, amounts of
adenine and thymine are always equal, and
amounts of guanine and cytosine are always
equal
4. James Watson and Francis Crick – published their work
showing the structure of DNA in 1953 (more detail later)
Names and Dates: Supplement to DNA Notes
Watson and Crick work together at the Cavendish Lab at Cambridge
University in England. Sir Lawrence Bragg was in charge at the Cavendish.
Watson and Crick take the approach of model building to solve the DNA
puzzle.
February 28, 1953: Crick announces in the English pub “The Eagle” that he
has found “the secret of life.” Watson and Crick published their work later
in 1953. Two other articles are published with their work, one of which is
Rosalind Franklin’s article. Her article is modified to make it appear that
her work was based on and supports Watson and Crick’s conclusions about
the structure of DNA. In reality, it’s the reverse – Watson and Crick’s work
is based on Franklin’s work that they deviously obtained through Maurice
Wilkins, Franklin’s colleague.
James Watson, Francis Crick and Maurice Wilkins received the Nobel prize
in 1962 for their work on DNA structure and how the DNA molecule can
function to carry genetic information.
Rosalind Franklin:
1. She worked at King’s College in London, England. The head of her
lab was J. T. Randall. Maurice Wilkins was a colleague of hers at
King’s. She and Wilkins often had personality clashes, partly
because Randall never clarified each one’s role at the lab.
2. She performed research on the DNA molecule using X-ray
crystallography to take pictures (she was one of the best
crystallographers in the world); this research was the basis of the
double helix shape to DNA that Watson and Crick are so famous for
discovering. Franklin took the approach of collecting data and then
analyzing the data for finding the structure of DNA. The idea of a
helical shape for DNA was all Rosalind’s. Without her work,
Watson and Crick would probably have not figured out the DNA
molecule before anyone else.
3. She died at age 37 in 1958, before Watson publishes his book and
before the Nobel Prize is awarded.
4. She was never acknowledged for her contribution to the structure of
DNA until Watson described her as a horrible person in his book
“Double Helix” (published in 1968). Everyone who was familiar with the
DNA story objected to Watson’s portrayal of her and the fact of
Rosalind’s work being so important was brought to the public’s
attention.
5. Beyond DNA research, Franklin contributed vast amounts of knowledge
to science in the areas of coal, viruses and their structure and function,
and even helped to create a far better gas mask for war torn Britain
during WWII. Her knowledge of X-ray diffraction would lead to her
travels throughout the US to give lectures at many of the most renowned
universities.
Maurice Wilkins:
Was the “assistant director” of the lab that Franklin worked in; he was good
friends with Crick and was most likely the conduit through which Franklin’s
work (including Photo 51) is given to Watson and Crick; received the Nobel
Prize with Watson and Crick in 1962.
Alfred Nobel:
Was a major business man in mid to late 1800’s; invented dynamite and made
his huge fortune from it; his brother died and a newspaper printed Alfred’s
obituary by accident and called him the “merchant of death” (dynamite killed
a lot of people in demolition accidents); Alfred hated the thought of his legacy
being such a terrible one so when he died he left most of his money for the
establishment of the Nobel Prizes; it’s the highest award a person can receive
and covers lots of different categories (literature, physics, medicine, peace...)
Erwin Chargaff:
1. He studied DNA and analyzed how much thymine, cytosine, adenine and
guanine were in each sample. He found that the amounts of thymine and
adenine were always equal, and the amounts for cytosine and guanine
were always equal.
2. The result was Chargaff’s Rule:
A = T and C = G
DNA Replication
I. Replication – process of copying DNA
A. The structure of DNA lends itself well to being copied.
1. Need to be able to copy DNA for cell division
and for reproduction
2. The accuracy of replication is impressive – only
about 1 error in every 10 billion nucleotides –
and is especially remarkable considering the
speed of the process. However, despite proteins
that check for damage in the DNA or mistakes
in the copying process, replication is not 100%
foolproof. And these mistakes, though rare,
means that genes are sometimes altered during
replication.
3. The rules of base pairing (A-T, C-G) factor
heavily into the process
4. Remember, replication occurs during the later
part of interphase.
B. Prokaryotes vs. Eukaryotes
Prokaryotes:
1. In most prokaryotes, there is a lot less DNA to
copy. Most have a single circular DNA
molecule, its single chromosome, located in the
cytoplasm.
2. Replication begins at a single point in the
chromosome and proceeds in two directions
until the entire chromosome is copied.
Eukaryotes:
1. Eukaryotic chromosomes are generally much
bigger than those of prokaryotes.
2. Replication in eukaryotes may begin at
hundreds or even thousands of places on the
DNA molecule, each called an origin of
replication (special sites of DNA with specific
sequences of nucleotides). At each origin,
replication will proceed in two directions.
3. As replication proceeds, these areas eventually
will meet up with each other. Multiple areas
being copied at once makes the process much
faster than having a single origin of replication.
C. How Replication Happens
1. The two strands of the double helix separate, or
unzip. This is done by a series of enzymes.
Remember that the base pairs (the rungs of
DNA) are held together by hydrogen bonds.
These are relatively weak compared to the
covalent bonds holding the sugars and
phosphates together (sides of the ladder of
DNA).
2. This unzipping allows two replication forks (areas
where the DNA is unzipped and new DNA is
forming) to form. Each side or strand of the
unzipped DNA will be a template for a new strand
to form.
3. New nucleotides are added to each side according
to the rules of base pairing (A with T, C with G).
These nucleotides are called “free nucleotides” and
are free floating in the nucleus. The enzyme called
DNA polymerase joins individual nucleotides to
the unzipped sides to form a new strand (or side)
of DNA, completing the helix. DNA polymerase
also “proofreads” the new strand of DNA to avoid
errors.
4. The sugars and phosphates on the sides covalently
bond together to form a new backbone.
D. Semiconservative Process
1. At the end of replication, there are two copies of
the DNA molecule.
2. In each DNA molecule, one strand (or half of the
molecule) is from the original DNA that we started
with. The other strand was made from free
nucleotides and is the “new” strand.
3. This model of copying, where the “copy” actually
contains half of the original molecule, is called the
semiconservative model. Containing an original
strand helps to prevent mistakes.
DNA Replication
I.
The Role of RNA
A. Despite finding the structure of DNA, it was not at all
clear how a gene actually works. To find this answer, a
lot more research and experimentation had to be done.
1. Part of the answer was provided when researchers
found that another nucleic acid, ribonucleic acid
or RNA, was involved in putting the genetic code
into action.
2. In decoding the information of DNA, part of the
base sequence of DNA is copied into a section of
RNA.
3. The RNA then has the instructions to direct the
next step, which is making proteins. Proteins help
to determine the characteristics of an organism
and are a major part of genes being expressed.
B. RNA and DNA
1. Both DNA and RNA are nucleic acids, but there
are several structural differences, summarized in
the chart below.
DNA
Double stranded
RNA
Single stranded
Base pairs:
A–T
C–G
Base pairs:
C–G
A–U
U=uracil; replaces thymine
Deoxyribose is the sugar
Ribose is the sugar
2. Comparing the roles of DNA to RNA: DNA is
like the complete set of blueprints to build a
shopping mall. RNA is a disposable copy of one
tiny section of those blueprints, like the
plumbing for the Starbucks inside the mall. As
a builder, you would never let someone borrow
your blueprints. They are too valuable.
Similarly, the DNA doesn’t leave the nucleus.
Instead, RNA is used to pass along the
information.
II. Types and Functions of RNA
A. There are many types of RNA, but the majority of
them are involved with one job – to make proteins.
1. RNA controls the assembly of amino acids into
proteins.
2. Recall that proteins are made from chains of
amino acids, like beads in a necklace. A single
chain of amino acids is a polypeptide. Proteins
can be made of a single polypeptide or many
polypeptide chains bonded together.
3. The properties of a protein are determined by
the order in which different amino acids are
joined together. (The order of the amino acids
is determined by the order of the nucleotides in
the DNA, but you’ll see that later.  )
4. Recall as well that proteins are made in the
ribosomes of the cell.
B. While there are other types, we will discuss the 3
main types of RNA used in protein synthesis (which
is the process of making proteins).
1. Ribosomal RNA (rRNA) – ribosomes are made
of 2 subunits and each subunit is composed of
several rRNA molecules and lots of different
proteins
2. Messenger RNA (mRNA) – used to send
information from the DNA to the ribosome;
serves as a disposable copy of the information in
the DNA
3. Transfer RNA (tRNA) – each tRNA molecule
has an amino acid attached to it; the tRNA
brings the amino acid to the ribosome – when
the tRNA and the mRNA match up, the amino
acid is added to the chain of amino acids
III. Protein Synthesis
A. Protein synthesis : process of making proteins
1. Most genes contain instructions for assembling
amino acids into proteins.
2. Proteins play a large role in determining an
organism’s characteristics.
3. RNA is integral to this process.
4. There are two parts to protein synthesis –
transcription and translation
B. Transcription
1. During transcription, a segment of DNA is
copied into a complementary strand of mRNA.
a. Requires the enzyme RNA Polymerase,
which binds to the DNA and separates the
strands.
b. RNA polymerase then uses one strand of
DNA as a template for the nucleotides being
assembled into a strand of mRNA.
2. How does the RNA polymerase know where to
start on the DNA?
a. The enzyme only binds to promoters.
Promoters are regions of the DNA that have
specific base sequences that signal RNA
polymerase exactly where to start making
mRNA.
b. Other signals result in transcription
stopping.
3. Transcription - Prokaryotes vs. Eukaryotes
a. In prokaryotes, the mRNA (made in the
cytoplasm) goes straight to the ribosome for
the next step in protein synthesis –
translation.
b. In eukaryotes, the mRNA (made in the
nucleus) is first spliced inside the nucleus.
i.
Splicing: removing the parts of
mRNA that aren’t needed
ii. Introns are the segments of mRNA
that are removed. Exons are the
segments that are spliced back
together. (exon  expressed)
iii. We’re not entirely sure of the reasons
behind splicing.
4. After splicing, the mRNA leaves the nucleus
and goes to the ribosome for the second part of
protein synthesis, translation.
Splicing is pictured above.
Splicing only occurs in eukaryotes.
Transcription is pictured above. Remember that in
prokaryotes, transcription happens in the
cytoplasm. In eukaryotes, it occurs in the nucleus.
Note that the mRNA is single stranded & has uracil,
while DNA is double stranded & has thymine.
C. Translation
1. Translation – converting the information in the
mRNA into a chain of amino acids (polypeptide
or protein)
2. How can a code of just 4 letters carry
instructions for 20 different amino acids?
a. The genetic code (in the form of mRNA) is
read three “letters” at a time, so that each
“word” is 3 bases long and corresponds to a
single amino acid.
b. Each three letter “word” in mRNA is
known as a codon.
i.
Codon: a 3 base section of mRNA
ii. Each codon specifies a single amino
acid that is to be added to the
polypeptide chain
c. How do you “read” the codons to find the
right amino acid? USE THE CHART!!!
d. Since there are 64 different combinations
of the 4 letters (43), most amino acids have
more than one codon that will code for it.
For example, how many codons are there
for the amino acid called Leucine?
e. There are also start and stop codons, which
act as punctuation marks of a sort.
i.
The start codon (AUG) is for
Methionine.
ii. How many “stop” codons are
there?
3. Steps of Translation
a. Begins when a ribosome attaches to an
mRNA molecule in the cytoplasm.
b. The mRNA is read one codon, or 3 bases, at
a time.
c. As each codon passes through the ribosome,
tRNA’s bring the proper amino acids to the
ribosome.
i. Each tRNA molecule carries just
one kind of amino acid.
ii. Each tRNA has 3 unpaired bases
sticking out. These 3 bases on the
tRNA are called the anticodon.
iii. Each tRNA is complementary to
(bonds with) one mRNA codon.
iv. Only when the anticodon and the
codon bond together is the amino
acid accepted and added to the
chain.
d. The ribosome helps a peptide bond to form
between the amino acids as they are added.
e. As each amino acid is removed from the
tRNA, the now “empty” tRNA’s are
released from the ribosome. They will
“recharge” with another of the same amino
acid in the cytoplasm to be used again.
TRANSLATION
B. The Polypeptide “Assembly Line”
The ribosome joins the two amino acids –
Methionine and phenylalanine – and breaks the
bond between methionine and its tRNA. The
tRNA floats away from the ribosome, allowing
the ribosome to bind another tRNA. The
ribosome moves along the mRNA, binding new
tRNA molecules and amino acids.
Remember: DNA  RNA  Protein!!!!