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DNA & Protein Synthesis
Chapter 18
The Molecule of Heredity
Seeking the Genetic Material
• 1928 Griffith finds that virulent bacteria
can transform nonvirulent bacteria into
the deadly form. Virulent: able to cause
disease Animation & Game
• 1944 Avery: found DNA was the
molecule of heredity, not protein or
RNA.
Avery, Macleod, McCarthy
• 1952 Hershey and Chase: found that
viruses injected DNA into host bacteria.
and Chase
DNA is confirmed as the unit of Hershey
Experiment
heredity.
Who Else Contributed?
• Chargaff: 1949 discovered there are
always equal amounts of Adenine to
Thymine and the same percentage of
Cytosine to Guanine.
Chargaff’s Ratios
• Rosalind Franklin 1950-53: Photographed
DNA with X-rays.(pg 187, Fig 10-4) Found
helictical shape (i.e. Helix Shape) while
working in the lab of Maurice Wilkins.
Franklin Click:
NOVA News Minute
16.7 The Structure of DNA
• Structure was discovered by Watson and
Crick in 1953, 1962 receive Nobel Prize.
• Double Helix:
• Nucleotides: Phosphate, Sugar
(deoxyribose) and a Nitrogen base
(A,T,C,G)
• DNA Backbone: Sugar-Phosphate-Base
Connected by Phosphodiester linkage
• 5 carbon sugar and phosphate molecule
are the same for each nucleotide.
Nitrogen base changes.
Deoxyribonucleic Acid (DNA
Continued)
• 2 strands one strand runs 5’ to 3’ the other
3’ to 5’ (upside down)
• Where does the 5’ and 3’ come from? Read
Synthesize
from 5’ to 3’
DNA
From
Here
Nitrogen Bases: 16.8
• Adenine double bonds to Thymine
• Cytosine triple bonds with Guanine
• GCAT Bases are held together by weak
hydrogen bonds.
• Purines (A&G) vs. Pyrimidines (C,T,U)
• A mistake here is a Point Mutation.
• Can you tell me the complementary
strand for : AATCGCGA?
______________
Eukaryotic DNA Replication
• Overview Cinema: The Forest from the
Trees
• The DNA Unzips, and each strand serves
as a template for the formation of a new
strand composed of complementary
nucleotides G-C, A-T
HHMI Replication
Replication: Entering the Forest
• DNA Replication begins at special Sites
called Origins of Replication
• Where the two strands of DNA split is
called replication bubbles (thousands)
• Why do you need thousands of these
bubbles? (Hint: The DNA molecule has 3
billion base pairs)
DNA Replication 16.12
• Double helix unwinds using DNA
Helicase.
• DNA Helicase breaks the hydrogen bonds.
• Where the DNA breaks apart is called the
replication fork. DNA polymerase
(another enzyme) adds nucleotides at the
5’ to 3’ end towards the replication fork.
• FYI: Humans= 50 nuclotides/sec
Meselson & Stahl
Experiment
DNA Replication Explained
What about the 3’ to 5’ end?
• Need RNA Primer. (RNA and the enzyme
primase).
• Lagging strand- away from the replication
fork replicates using Okazaki fragments.
• Each 100-2000 nucleotide Okazaki
fragment is joined by DNA ligase.
• Fig 16.15 & 16.16
Checking for errors
• DNA Polymerase also proof reads the
strands- Mismatch Repair
• A mistake in nucleotide pairing is a
Mutation
• Multiple replication forks happen all at
once so that the process is speedy.
DNA Review
Flashcards
How do we keep the replication
process clean?
• Single-stranded binding proteins scaffold the two
DNA strands apart.
• Topoisomerase cuts DNA that is wound too tightly.
• Telomeres, made by the enzyme telomerase
protect against gene loss at the end of the
chromosome. Fig 16.19
– Telomeres repeating sequences of TTAGGG (thousands
of times)
– Telomeres shorten with each cell division. Aging
anyone?
How do bacteria replicate their
DNA? Fig. 16.2
The Technical
Details
The Non-Technical
Details
The Technical
Details
What is RNA and how is it useful?
• RNA= Ribonucleic Acid
• Single stranded. Ribose sugar.
• Transcribes DNA and Translates it into
proteins.
• Proteins are organic compounds that have
specific jobs in the cell. (Ex. Enzymes)
PBS Video
RNAi
What are the 3 types of RNA?
• mRNA= Messenger RNA
– Transcribes or rewrites DNA’s message as
mRNA, mRNA carries message to ribosome
• rRNA = Ribosomal RNA. Fig. 17.16
– Creates the ribosomes on the rough ER and
cytoplasm where proteins are made.
• tRNA = Transfer RNA. Fig. 17.14 & 17.15
– Transfers amino acids to the ribosomes and
translates the mRNA into protein. (Called
Translation because the message changes
from nucleic acid to protein, a different
organic compound)
Transcription and Translation
Cinema Fig. 17.3
• Don’t relax too much
• Pencils out? Notes ready? Lets work!
Overview Movie
Start Here
HHMi Transcription
More Detail
AUG (start)
methionine
First
Base
U
C
UAA UGA UAG (stop)
Second Base
Third
Base
U
C
A
G
UUU
phenylalanine
UCU
serine
UAU
tyrosine
UGU
cysteine
U
UUC
phenylalanine
UCC
serine
UAC
tyrosine
UGC
cysteine
C
UUA
leucine
UCA
serine
UAA
stop
UGA
stop
A
UUG
leucine
UCG
serine
UAG
stop
UGG
tryptophan
G
CUU
leucine
CCU
proline
CAU
histidine
CGU
arginine
U
CUC
leucine
CCC
proline
CAC
histidine
CGC
arginine
C
CUA
leucine
CCA
proline
CAA
glutamine
CGA
arginine
A
CUG
leucine
CCG
proline
CAG
glutamine
CGG
arginine
G
What is Wobble?
• A relaxation of the base pairing rule.
– Example: UCU, UCC, UCA, UCG = serine
• Why is their Wobble room?
– Allows for point mutations not to cause an
incorrect protein to be made.
– Since there are only 20 Amino Acids and 64
codons to code for them we can have
repeats.
What are the three stages of
transcription?
• Initiation: RNA polymerase binds to the DNA
promoter. Fig 17.8
– TATA box is the area that transcription factors
recognize within the promoter.
– Creates a transcription initiation complex
(transcription factors & RNA polymerase bond on
promoter)
• Elongation: DNA template continues to grow as
RNA polymerase separates DNA strand.
– Transcribed part: Transcription unit (made of codons)
(1 gene transcribed many times in caravan fashion)
• Termination: mRNA is cut free from DNA template
•
•
•
•
•
How do Eukaryotes control
Gene expression?
Transcription creates Pre-mRNA. Fig 17.10
Pre-mRNA includes Introns and Exons
Introns= Fillers (intervening sequences)
Exons= Code for proteins (expressed sequences)
mRNA is just the Exons
Eukaryotic Transcription
How else is mRNA processed for
translation?
• snRNP’s (“snurps”) & splicesomes remove
introns, creating the final mRNA sequence=
Exons only. Fig. 17.11
• What stops the mRNA from degrading by
hydrolytic enzymes & binds it to the
ribosome?
– 5’ cap: modified guanine nucleotide is added to
the 5’ end.
– A poly tail: 30-200 adenine nucleotides on the
3’ end. (Also helps release mRNA into the
cytoplasm.
What do enhancers do? Fig 18.9
• Enhancer’s causes the gene it is
enhancing to be expressed. Transcription
occurs in eukaryotes when an enhancer
site is activated.
Fig. 18-9-3
Promoter
Activators
DNA
Enhancer
Distal control
element
Gene
TATA
box
General
transcription
factors
DNA-bending
protein
Group of
mediator proteins
HHMI Transcription
Basic
RNA
polymerase II
RNA
polymerase II
Transcription
initiation complex
RNA synthesis
Fig. 18-8-3
Putting it all together
Enhancer
(distal control elements)
Poly-A signal
sequence
Termination
region
Proximal
control elements
Exon
Intron
Exon
Intron Exon
DNA
Upstream
Downstream
Promoter
Primary RNA
5
transcript
Transcription
Exon
Intron
Exon
Intron Exon
RNA processing
Cleaved 3 end
of primary
transcript
Poly-A
signal
Intron RNA
Coding segment
mRNA
3
5 Cap
5 UTR
Start
codon
Stop
codon
3 UTR Poly-A
tail
And now Translation Fig. 17.17-17.19
• Translation: mRNA codons code for an Amino
acid sequence.
• tRNA attaches to the ribosomal rRNA with
anticodon. GTP recycles tRNa with enzyme.
• Ribosome: protein and rRNA created in the
nucleolus.
– Bacterial & Eukaryotic ribosomes differ, this allows
Tetracycline and Streptomycin to stop protein
production in bacterial ribosomes.
– Small and large subunits for attachment to mRNA.
– Three binding sites for tRNA
• P site: Holds the growing polypeptide chain
• A site: Holds the newest tRNA
• E site: Exit site
What are the three stages of
translation?
• Initiation: mRNA’s AUG codon (methionine)
attaches to the ribosome. tRNA attaches.
• Elongation: forms the polypeptide chain as
tRNA’s continue to attach.
• Termination: mRNA reaches a stop codon.
Release factor hydrolyzes the bond. mRNA is
degraded. Polypeptide is freed from ribosome.
– After this the polypeptide will fold or pleat (secondary
structure), Chaperonine will complete the tertiary
structure.
How Proteins fold
Fold it, the Game!!!
What about when a mistake
occurs?
• They can effect the species gene pool.
• Point mutation= single base-pair
substitution. (sickle cell)
• Insertion or deletion= frameshift
mutations. The Dog -> HeD og
• Missence Mutation= If a codon within a
gene turns into a stop codon translation is
altered.
Gel Electrophoresis
• How does this link to Gel Electrophoresis?
• Below is a reminder of what
Electrophoresis is and how it works.
Gel Electrophoresis