Transcript DNA replication and protein synthesis

DISCOVERY OF DNA
PROTEIN SYNTHESIS
CHAPTER 10
Discovery of DNA
• Discovery of genetic material can be
traced back to the late 1920’s
• Many scientists help with the
understanding of the function and
structure of DNA
I. DISCOVERY OF DNA
A. Frederick Griffith
Trying to discover a vaccine for pneumonia
– Experiments with bacteria causing pneumonia
– 2 strains
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S - caused disease (virulent)
R - nondisease
B. Griffith’s Experiment
• Exp. 1
– Injected R into mice - they lived
• Experiment 2
– Injected S into mice - died
• Experiment 3
– Heated S strain (to kill it, but not the
mucous coat) and injected it – Result - lived
Griffith’s Exp. Cont…
• Exp. 4
– Combined R strain with heated S strain
– Hypothesis - harmless
– Results - died of pneumonia
• Conclusion:
– Some material passed from dead S strain to live R
strain changing R strain into S strain.
– He called this transformation (process bacteria
change by absorbing genetic material from outside
source.
C. Oswald Avery
• American
• Wanted to figure out what transforming
agent was
• Hypothesized - DNA RNA or protein
Avery Experiments
• Experiment 1
– Protease - destroys protein in heat killed S
strain
– Added protease to heated S strain, then
mixed with live R strain
– Result = all died
– Therefore PROTEIN is not the
transforming factor.
Experiment 2
– Used RNase - destroys RNA
– Mixed RNase with heated S strain then
with R strain - mice died
– Concluded - RNA not transforming factor
Experiment 3
• Used DNase to destroy DNA
• Mixed DNase with heated S then with live R
injected into mice
• Result - all lived
• Concluded - DNA was the transforming
factor. Cells missing DNA did not transform
R strain into S strain and therefore mice lived
D. Hershey and Chase
• Alfred Hershey & Martha Chase
• American
• Used viruses to test whether DNA or
protein was transforming factor when
viruses enter bacteria
Hershey and Chase cont
• Viruses that infect bacteria =
bacteriophages (phages)
• Viruses infect and destroy bacteria
• Viruses only contain protein and DNA
• Bacteriophages only contain protein
coat and DNA
• So can use these to determine if protein
or DNA is the transforming factor
Experiment 1
• Labeled protein of T2 virus with radioactive
sulfur
• Allowed virus to enter bacteria (creating
bacteriophage)
• Put this in blender and centrifuged it to
separate the bacteria from virus
Results - found NO radioactive sulfur in the
bacterial cell
Concluded - protein was not the transforming
factor
Experiment 2
• Labeled DNA of T2 virus with radioactive
phosphorus
• Allowed virus to enter bacteria
• Put this in blender and centrifuged it to
separate bacteria from virus
• Results- found radioactive phosphorus
entered bacterial cell
• Concluded- DNA is the transforming factor
Pauling, Wilkins,Franklin
• At this point most biologists were
conviced that DNA was genetic
material, but were uncertain of the
structure.
• Used X-ray crystallography image of
DNA
• Watson and Crick used this x-ray to
summize shape of DNA
•
Watson and Crick
• Utilized many scientists ideas to create
first 3-D DNA model
• Franklin,Pauling, Wilkins
• Erwin Chargaff - complementary base
pairing - figured out A-T and C-G
Structure of DNA
• Structure of deoxyribonucleic acid
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Double helix (twisted ladder)
5 Carbon sugar (deoxyribose)
Phosphate
Nitrogen base (A, T C, G)
The 5 carbon sugar and phosphate are the sides of the
ladder (covalently bonded together - strong bond) and
the nitrogen bases are the rungs.
DNA
DNA
DNA Nitrogen Bases
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Complementary bases
Adenine pairs with Thymine
Cytosine pairs with Guanine
Nitrogen bases held together by weak
hydrogen bonds
• Adenine and guanine larger - purines
• Cytosine and thymine smaller - pyrmidines
Purines and Pyrmidines
DNA Replication
• Semi-conservative model – uses a strand (
half) of its DNA to make a new strand of
DNA
• Occurs S phase of interphase
• Occurs to assure that each cell going
through cell division has the identical DNA
so can perform identical function.
Enzymes of DNA Replication
• DNA polymerase –
• There are few different types of this
• Attaches complementary bases to template strand
• DNA ligase– Joins Okazaki fragments (O-kah-zocki) together
• Primase– Creates “starting point” for nucleotides to be added onto
strand
• Helicase
– Enzyme that breaks “weak bonds” and unwinds DNA
• Proteins
– Other enzymes
Steps of DNA Replication
1. Helicase breaks bonds, unwinds and unzips DNA.
This creates a replication bubble/ fork
2. Helicase separates DNA creating a
replication form.
•
Creates a leading and lagging strand of
the DNA
• Binding proteins prevent strands from
reattaching to each other
3. Leading Strand
• RNA primer attaches to leading strand to give it a
“starting point”
• DNA polymerase adds complementary nucleotides
(A-T and C-G) from 3 to 5 prime end of leading
parent strand (so is really creating the opposite or a 5
prime strand)
• The DNA polymerase can only add bases to the 3’
end of the strand, never to the 5’end
So really creating a 5-3 strand (opposite of the leading
strand template)
*** 3’ end refers to where the OH is attached to the
deoxyribose (attached to 3rd carbon) while 5’ end
the phophate group is attached to the 5th carbon.
• Adds bases in direction toward replication fork.
4. Lagging Strand
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RNA primer attaches to strand close to the replication
fork to create a starting point (3 end of the template
lagging strand)
DNA polymerase attaches complementary bases
beginning at the RNA primer – adding bases toward the
5 end of the strand (away from the replication fork)
This creates okazaki fragments.
•
Creating an opposite strand from the template or a 3 – 5
strand
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When completed, RNA primase is replaced by DNA and
ligase bonds all DNA together.
•
5. DNA binding proteins then go through each
strand and correct any mistakes on DNA
strands “cleans it up”
6. Two Identical strands of DNA
Semi-conservative model - because it uses
1/2 of strand for a template to create 2
strands of DNA
• http://www.stolaf.edu/people/giannini/fla
shanimat/molgenetics/dna-rna2.swf
PROTEIN SYNTHESIS
• Process of using DNA to make protein
• Function of protein in body
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Enzymes
Immunity
Communication between cells
Build muscles
Characteristics (skin color, eye color, etc…)
Structure of DNA
• Double helix
• Nucleotides
– Dexyribose (5 carbon sugar)
– Phosphate
– Nitrogen base (A,T,C,G)
DNA
DNA
BASES
FUNCTION OF DNA
• Contains genetic information
• Recipe to make any and all protein your
body needs to function everyday
HELPERS
• RNA - ribonucleic acid
– Acts as a messenger between DNA and
ribosomes
Ribosomes - where protein is “constructed”
3 types of RNA
mRNA
tRNA
rRNA
3 Stages of Protein synthesis
• 1. Transcription - “write recipe down”
• 2. Translation - “put recipe together”
• 3. Elongation - “glue” amino acids
together to form protein
SUMMARY
Transcription
• Occurs in nucleus of cell
• Process by which molecule of DNA is
copied into complementary strand of
mRNA
Steps of Transcription
1. DNA unwinds and unzips (helicase)
(only in area of the recipe) creating 2
strands. An active strand and
“dummy”strand
2. Active strand is the one to be used to
make the protein (the template)
3. Special sequence of DNA is
recognized by RNA polymerase as the
“start signal” (promoter)
Steps of transcription cont…
4. RNA polymerase matches up complementary
bases between DNA and RNA (A-U, C-G),
using DNA as a template
5. RNA polymerase moves along the area of
the DNA with the recipe, matching up
complementary bases
6. when it hits the “stop codon” mRNA drops
off DNA
7. 7. At this point mRNA has “copied” the
recipe for the protein.
8. DNA winds back up and mRNA gets
modified before leaving the nucleus
TRANSCRIPTION
mRNA modification
• While still in the nucleus, mRNA gets
modified.
• mRNA consists of exons and introns
• An enzyme comes along and splices out the
introns (pieces of DNA) that is not part of the
recipe needed for the protein.
• Exons are then spliced together to create the
“real recipe” for the protein your body needs
• Exons are capped and tailed for protection
and then leave the nucleus via nuclear pores.
EXONS AND INTRONS
WHY COPY DNA????
• DNA is too large to leave the nucleus,
so it needs a messenger to bring
genetic information to the ribosome in
the cytoplasm.
• This messenger is the mRNA
TRANSLATION
• Process of decoding mRNA into protein
• Code being translated from language of
nucleic acids into polypeptide
STEPS OF TRANSLATION
• 1. mRNA attaches onto a ribosome and the
first codon “start” is read by the ribosome.
This signals ribosome to start translating the
recipe
• 2. The ribosome reads each codon of mRNA
and signals tRNA (complementary nitrogen
bases which are carrying a specific amino
acid). (Also called an anticodon)
• 3. Complementary tRNA matches up with
mRNA codon, and brings the amino acid
along with it.
IN CYTOPLASM
Translation cont….
• 4. The ribosome moves along the mRNA
reading it and signaling tRNA to bring amino
acids to the ribosome.
• This continues until the ribosome hits the stop
codon.
• 5. When hit stop codon, mRNa breaks off
and returns to the nucleus (disassembles)
• 6. All that is left is a string of amino acids in a
specific order. This specific order is what
determines the name and type of protein that
was just made.
TRANSLATION ON
RIBOSOME
TRANSLATION
ELONGATION
• The string of amino acids is bonded
together (during the process of
translation) to create the protein that
your body needs.
TRANSLATION
MUTATION
• What will occur if the amino acids are
not in the right order?
• What happens if you did not eat enough
protein in your diet and the tRNA could
not pick up a specific amino acid
needed to make the particular
protein???
ROLE OF GENE
EXPRESSION
• “Turning on “ of a gene that results
from transcription and translation
• By regulating gene expression, cells are
able to control which part of genome will
be expressed.
GENE EXPRESSION IN
DEVELOPMENT
• Every cell in developing zygote contains
all of the organism’s genes, only small
number of genes expressed
• Certain genes are turned on and off as
proteins are needed at different times
during the organism’s life
Development
• As organisms grow from zygote the cells
differentiate
• Differentiate - cells develop into different
types of cells and tissue.
• Ex. Nervous cells, skin cells, muscle, etc…
• The development of form in an organisms is
called morphogenesis.
Morphogenesis
• Homeotic genes regulate where certain
anatomical structures go.
• Ex. Appendages, organs, etc…
• Homeotic gene is the master genes of
development and determine how the body will
be organized
• These genes regulate development by
switching genes on and off, which controls
the rate of cell division in certain areas of the
developing organism, this results in specific
patterns of structural development
Changes in Gene Expression
• Control of gene expression is also
important throughout life.
• Cells constantly switch genes on and off
to express characteristics
Gene Expression and Cancer
• Proto-oncogenes - regulate cell growth, cell
division and the ability of cells to stick
together.
• Ensure that mitosis runs smoothly
• A mutation that leads to over expression of
proteins that control mitosis can effect the cell
cycle and lead to cancer.
• Some genes act as Tumor suppressor genes
that prevent cell division from occurring too
often.
• In cancer tumor suppression genes are
damaged
Kinds of Cancers
• Carcinomas - cancer in skin and tissues
lining organs of body
– Ex. Lung cancer
Sarcoma - cancer in bone and muscle
Lymphoma - tumors in blood forming tissues
Ex. leukemia