Download DNA History, Structure, and Replication

DNA History, Structure, and Replication
QUESTION:
Where is genetic information stored?*
a)
b)
c)
d)
in the ribosomes of cells
within the proteins of cells
within the DNA of cells
within the membrane of cells
*But scientists did not always know this!
DNA is a very small molecule and had to
be discovered. Let’s look at the history,
structure, and replication of DNA in today’s
discussion.
The answer is here
somewhere!
Discovering DNA -
Once Mendel understood that “factors” could be
passed to offspring, scientists began to wonder
what these factors were. They were sure of a few
things…
1.The factors needed to be able to store information
2. The factors needed to be replicable
3. And the factors needed to sometimes undergo
changes (mutations)
1869 – Discovering Nucleic Acids
Swiss Physician, Johannes Friedrich
Miescher isolated a new biomolecule
he called “nuclein” from the nuclei of
white blood cells. This later became
called nucleic acids, but to be honest
– we had no idea what it did. We just
knew it was there.
Miescher used pus and bloodstained
bandages from a hospital to study
“nuclein”
Chemistry of NUCLEOTIDES (Nucleic Acid Monomers)
Analysis of the nucleic acid showed that it
contained a sugar, phosphate and one of four
nitrogen bases
Adenine
Guanine
Cytosine
Thymine
- Now we know what’s in it, but
what does it look like? What does it
do?
**THE BIG QUESTIONS**
Were these new nucleic acids the secret to
Mendel’s factors? Or were the everpresent proteins the secret? Where is the
cell’s genetic code?
a) Proteins contain 20 amino acids that can be
organized in countless ways to determine traits
b) Nucleic acids only contained 4 different nucleotides
Thanks to their 20 amino
acids, scientists were
leaning toward proteins.
As a information storage
option, amino acids would
have a lot of options, since
you could combine these in
different ways
The 4 bases of the NAs
didn’t seem to have as much
versatility.
It’s like have an alphabet of
20 letters versus and
alphabet of 4.
The Experiment that finally gave us an answer…
Frederick Griffith was attempting
to find a vaccine against
pneumococcus bacteria that
caused pneumonia. In this
experiment, he accidentally
determined the true source of
the genetic code by discovering
that one type of bacteria could
actually turn into another. Let’s
take a look at his work…
An overview of the transformation experiment.
Can you summarize this in your own words?
DNA was determined to be the genetic code.
● DNA from the dead S strain bacteria was taken in by the
live R strain, causing them to transform into S strains.
● Denaturing the proteins or using enzymes that stopped S
proteins did not stop the transformation
● Using enzymes that denatured the nucleic acid did stop
the transformation
● We knew the DNA contained the instructions, but we still
didn’t understand how… All that information transferred
using only four “letters”?! How is that possible?!
Quick Recap:
1) What caused scientists to believe that proteins contained the
genetic code?
2) What was Miescher’s contribution to genetic studies?
3) How were the nonvirulent R strain bacteria transformed into a
virulent strain?
4) Griffith’s experiment resulted in which conclusion?
Alfred Hershey and Martha Chase Experiments
The bacteriophage – a virus used for studying DNA
Reminder: Viruses infect by injecting their DNA into a cell and taking
control of the host. Two types– Lytic viruses immediately use the host to
replicate their own DNA, make more viruses, and then lyse the cell to
release offspring. Lysogenic viruses actually insert their DNA into the
genome of the host. They go lytic eventually, but for a while, they “hide”
in the host DNA and become a part of the host organism. The
bacteriophage is a lysogenic virus for bacteria. How could we use them?
Bacteriophages
–
viruses that infect bacteria
●Using the bacteriophage to prove DNA as
the genetic code
● Bacteriophages have a protein capsid surrounding a piece
of DNA
● Experiments used radioactive sulfur to tag proteins and
radioactive phosphorous to tag the DNA.
● The goal was to see which substance (protein or DNA)
moved into the infected cell
Conclusion: The radioactive tag on the DNA went into the bacteria, not
the tagged proteins
But we still had NO clue what it looked like!
Imagine you had all of the pieces to a puzzle but you didn’t
know how they fit together. Scientists had the pieces of DNA.
Fame and fortune would go to the one who solved this puzzle...
The Race to Establish the Structure of DNA
THE PLAYERS
THE PIECES
adenine
guanine
cytosine
thymine
deoxyribose
phosphate
Examine the data below. Do you notice a pattern?
So did Erwin Chargaff...
Chargaff’s Rule
Amount of A, T, G, C varies by
species, but #A = #T and #G = #C
(#Purines = #Pyrimidines)
all species
had similar
ratios of
A, T, G, C
Purines (A&G) have two rings on the nitrogen base
Pyrimidines (C&T) have one ring on the nitrogen base
Could it be that
these pieces of
the puzzle fit
together…..
But what about
deoxyribose and
phosphates,
where do those
pieces fit? And
what is the
shape of DNA?
ROSALIND FRANKLIN & WILKENS
Took pictures of DNA structure with X-RAY DIFFRACTION
These
images
provided
clues to
the shape
of the
DNA
molecule.
Competition in science was a major theme during this period of time.
(1940-1953 ish)
Scientists often wanted to get sole credit for a discovery, and were
reluctant to share results with others. None of the work by Chargaff,
Franklin, and Wilkins were shared, they existed as isolated facts.
For consideration…..
1) Do you think scientists today are more likely to collaborate?
2) How has the internet changed science?
Enter……...WATSON & CRICK
They were criticized for their
methods which included
- hanging out in pubs and
talking about stuff
- playing cricket
- stealing data from other
scientists (Franklin/Wilkins)
- But they used Chargaff’s
rules and F/W’s picture to
mathematically determine
the double helix shape of
DNA. They won the Nobel
Prize for “solving the
puzzle” and mostly- they get
the credit. Fair?
DNA:
DOUBLE HELIX (Twisted Ladder)
Steps of ladder are bases (A, T,
G, C)
Sides of ladder are sugar &
phosphate
Sugar and phosphate are
covalently bonded along the
sides (strong!) while the steps
are hydrogen bonded (weak.)
5' and 3' ends
5'
4'
1'
3'
2'
2'
3'
4'
1'
Each side is antiparallel (runs in the
opposite direction). The numbers used to
represent each side refer to the carbons
attached to the deoxyribose and
covalently bonded to the phosphate. If
it’s connected at the 3rd Carbon, it’s the
3’ end. Vice versa for 5’.
5'
5’ and 3’ ENDS
Each Side is ANTIPARALLEL
Nucleotide =
o1 base
oDeoxyribose (sugar)
o1 phosphate
DNA is a polymer made
from nucleotide
monomers
What’s wrong with
this drawing?
DNA REPLICATION
-the process by which DNA
makes a copy of itself during S
phase of interphase prior to
ANY cell division
-replication is semiconservative, because one half
of the original strand is always
saved, or "conserved” in the
new strands
Looking at the pic, explain
replication in your words.
DNA Replication Steps
1. Protein enzyme(s) DNA helicase unzips the hydrogen bonds and
creates replication forks at several places along the molecule. DNA
binding proteins hold the separated strands apart to prevent
reannealing (reattaching).
2. Protein enzyme Primase creates a small section of nucleotides to
which protein enzymes DNA polymerases can fully attach and run down
the strand adding free nucleotides (following base pair rules) to the
exposed strand, covalently binding the sugars and phosphates on the
new side, and proofreading/correcting mistakes along the way.
***DNA polymerase can only travel down the strand from the 3' to the 5' end. Careful:
it only “reads” the OLD strand from 3-5, meaning it “builds” the NEW strand from 5-3.
What issue does this create?***
5. Since one side of the DNA runs in the 3’ to 5’ direction, it
is copied continuously and called the leading strand. The
other side runs in the 5' to 3' direction and is called the
lagging strand. Since the DNA polymerase can only READ
from 3’ to 5’ and BUILD from 5’ to 3’, this lagging strand
must be done in chunks called OKAZAKI FRAGMENTS.
Primase places a primer periodically, and DNA Polymerase
“jumps” from primer to primer working in reverse. These
fragments are eventually connected by another protein
enzyme called DNA LIGASE
6. Using multiple replication forks (sometimes called
replication bubbles) all down the strand, these enzymes
are able to copy all of the DNA relatively quickly.
DNA Replication
Which side is
leading? Which
side is lagging?
Which side will
be made
continuously?
Which side will
be made in
Okasaki
fragments?
Pg 235
Figure 13Ac