Download DNA: Structure, Function, and History

DNA: Structure,
Function, and History
The Nucleus
Nuclear envelope
- separates cytoplasm from the
nucleus
Nuclear
envelope
Nuclear
pore
Nuclear pore
- controls movements in and
out of the nucleus
Chromatin (DNA)
- genetic information
Nucleolus
- produces ribosomes
Nucleolus
Chromatin
Terms Used in DNA Science – a
cookbook analogy
„
Genome – all of the DNA of an organism
‰
„
Chromosomes – segments of DNA
‰
„
Chapters in the DNA cookbook.
Genes – individual genetic instructions on the
chromosomes
‰
„
The entire Cookbook
Recipes in the cookbook chapters
Alleles – variations of a gene
‰
Different or altered recipe for the same food
Miescher Discovered DNA
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1868
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Johann Miescher investigated the chemical composition of
the nucleus
„
Isolated an organic acid that was high in phosphorus
„
He called it nuclein
„
Today, we call it DNA (deoxyribonucleic acid)
Griffith Discovers Transformation
„
1928
„
He was attempting to develop a vaccine
„
Isolated two strains of Streptococcus pneumoniae
bacteria
‰
Rough (R) strain was harmless
‰
Smooth (S) strain was pathogenic and lethal
Griffith Discovers Transformation
1. Mice injected with
live cells of harmless
strain R
2. Mice injected with live
cells of killer strain S
3. Mice injected with
heat-killed S cells
4. Mice injected with
live R cells plus heatkilled S cells
Mice live. No live R
cells in their blood
Mice die. Live S cells in
their blood
Mice live. No live S cells
in their blood
Mice die. Live S cells in
their blood
Transformation
„
„
„
What happened in the fourth experiment?
The harmless R cells had been transformed by
material from the dead S cells
Descendents of the transformed cells were also
pathogenic (disease causing)
What Is the
Transforming Material?
„
„
„
„
In 1944, Avery found that cell extracts treated with
protein-digesting enzymes could still transform
bacteria
Cell extracts treated with DNA-digesting enzymes
lost their transforming ability
Concluded that DNA, not protein, transforms
bacteria
Later, Hershey and Chase also determined that
DNA was the molecule of genetic inheritance.
Hershey – Chase Experiment
Martha Chase and Alfred Hershey in 1953
2nm diameter overall
Structure of
DNA
In 1953, James
Watson and
Francis Crick
showed that DNA
is a double helix
(but they had help
along the way!)
0.34 nm between
each pair of bases
3.4 nm length of each
full twist of helix
Timeline of DNA Discoveries
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„
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1860: Gregor Mendel describes the nature of a genetic component
in hereditary.
1869: Miescher discovers an acidic compound in the nucleus of
cells. He calls it “nuclein”.
1928: Griffith concludes that there is a genetic, heritable compound.
1929: Levene discovers that DNA is composed of deoxyribose
sugar, phosphates and nitrogenous bases.
1944: Avery, MacLeod, and McCarty discover that DNA is the
“transforming factor” Griffith discovered.
1949: Chargraff determines A & T are in same proportions and G &
C are in the same proportions.
1952: Hershey Chase Experiment – DNA found to be the
hereditary molecule
1953: Watson & Crick discover the structure of DNA
1966: Genetic Code discovered.
Watson & Crick: How’d They Do It?
„
Used Chargraff’s rule of paring
‰
„
„
A with T; G with C
Determined phosphates must
be on the outside with bases on
the inside
Used x-ray crystallography to
determine helical structure
Structure of Nucleic Acids
„
DNA and RNA are nucleic acids
„
Nucleic acids are polymers of nucleotides
joined to each other in a line.
„
The sugar of 1 nucleotide joins to phosphate
group of the previous nucleotide
„
Forms a sugar-phosphate backbone.
Structure of Nucleotides
in DNA
„
„
Each nucleotide consists of
‰
Deoxyribose (5-carbon sugar)
‰
Phosphate group
‰
A nitrogen-containing base
There are four bases:
‰
Adenine, Guanine, Thymine, Cytosine
„
A
G
T
C
Nucleotide Bases in DNA
The purines - “double-ring”
ADENINE
(A)
phosphate
group
GUANINE
(G)
deoxyribose
The pyrimidines - “single-ring”
THYMINE
(T)
CYTOSINE
(C)
Composition of DNA
„
Amount of adenine relative to guanine differs among
species
„
Amount of adenine always equals amount of
thymine, and amount of guanine always equals
amount of cytosine
„
A base pairing rule ; borrowed from another scientist
named Chargraff (he discovered the ratios, but did
nothing with them)
A=T and G=C
Watson-Crick Model
„
DNA consists of two nucleotide strands
„
Strands run in opposite directions
‰
„
5’ ----> 3’ paired with 3’----->5’
Strands are held together by hydrogen bonds
between bases
„
A binds with T (uses 2 hydrogen bonds) and C
with G (uses 3 hydrogen bonds)
„
Molecule is locked in a double helix shape
2nm diameter overall
Structure of
DNA
The Sugarphosphate
backbone
The appropriate
Nitrogen bases
pair in the middle
of the helix
0.34 nm between
each pair of bases
3.4 nm length of each
full twist of helix
Rosalind Franklin’s Work
„
„
„
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An expert in x-ray crystallography
Used this technique to examine
DNA fibers
Concluded that DNA was some
sort of helix
Watson and Crick “borrowed” her data (without
giving her much credit) for their model of DNA
structure
Animation of DNA structure
Rosalind Franklin
X-ray crystallographer that
produced image Watson
and Crick used to
determine helical structure
with two strands
The Double stranded DNA Structure
Helps Explain How It Duplicates
„
DNA is two nucleotide strands held together by
hydrogen bonds
„
Hydrogen bonds between two strands are easily broken
under correct conditions, creating two single stranded
“sides”
„
Each single strand then serves as template for a new
strand’s production, using base-pairing rules.
DNA
Replication
„
Each parent strand
“unzips” to guide the
production of new
DNA strands
„
Every New DNA
molecule is half “old”
and half “new”
new
old old
new
Base Pairing
during Replication
Each old strand
serves as the
template for a
complementary
new strand
Complementary – the
appropriate base-paired
nucleotide
Enzymes in Replication
„
Enzymes unwind the two strands and their
complementary base pairs unzip
„
DNA polymerase attaches new complementary
nucleotides
„
DNA ligase fills in gaps
„
Enzymes wind two strands together
DNA Repair-enzymes fix mistakes before
problems arise.
„
Mistakes can occur during replication
„
DNA polymerase can read correct sequence from
complementary strand and, together with DNA
ligase, can repair mistakes in incorrect strand
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DNA replication animation
How does a Sunburn Affect your DNA?
„
DNA can be damaged by
UV light.
„
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The enzymes and proteins
involved in replication can
repair the
damage…hopefully!
If not repaired, dimers can
cause cancer.
Pyrimidine dimer in
DNA
Goodsell, D. S. Oncologist 2001;6:298-299
Steps from DNA to Proteins
Same two steps produce all proteins:
1) DNA is transcribed to form RNA
‰
‰
Occurs in the nucleus
RNA moves into cytoplasm
2) RNA is translated to form polypeptide chains,
which fold to form proteins
Three Classes of RNAs made by
Transcription.
„
Messenger RNA
‰
„
Ribosomal RNA
‰
„
Carries protein-building instruction
Major component of ribosomes
Transfer RNA
‰
Delivers amino acids to ribosomes
A Nucleotide Subunit of RNA
uracil (base)
phosphate
group
ribose
(sugar)
This
Nitrogen
Base only
Found in
RNA –
Never
Found in
DNA!
Base Pairing during Transcription
DNA
G
C
A
T
RNA
G
C
DNA
C
G
T
A
DNA
C
G T
base pairing in DNA replication
A U
A
base pairing in transcription
Transcription – Making an RNA strand
from a DNA template strand
„
Like DNA replication…
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Nucleotides added in one direction
Unlike DNA replication…
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Only small stretch is template (reads the gene)
‰
RNA polymerase catalyzes
nucleotide addition
‰
Product is a single strand of RNA
Promoter – a Transcriptional Road sign.
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A base sequence in the DNA that signals
the start of a gene
For transcription to occur, RNA
polymerase must first bind to a promoter
Gene Transcription
transcribed DNA
winds up again
DNA to be
transcribed unwinds
mRNA
transcript
RNA polymerase
Adding Nucleotides
5’
growing RNA transcript
3’
DNA
3’
direction of transcription
5’
Transcript Modification
unit of transcription in a DNA strand
3’
exon
intron
exon
transcription
intron
5’
exon
into pre-mRNA
poly-A
tail
3’
cap
5’
snipped
out
snipped
out
Exons-information
That is kept!
5’
Introns- junk
Information
that is removed
3’
mature mRNA transcript
Transcription Animation
Translation – making a protein using
mRNA information
„
Uses a segment of mRNA (encoding the correct genetic
information to make a protein)
„
Ribosomes “decipher” the information contained in a
nucleotide strand and translate it into a strand of amino
acids (a protein) using tRNA molecules to help bring in the
correct amino acid at the correct time.
„
All of your proteins are made this way!
Genetic Code
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Set of 64 base triplets
„
Codons – a series of 3
nucleotides in a row in
mRNA
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61 specify amino acids
(only 20 amino
acids=repeats)
„
3 stop translation
„
Directs a mRNA sequence
into a protein sequence
tRNA Structure
At lease one tRNA for
Each amino acid to be
Brought to the ribosome.
codon in mRNA
anticodon
amino-acid
attachment site
amino
acid
OH
Ribosomes
tunnel
small ribosomal subunit large ribosomal subunit
intact ribosome
Three Stages of Translation
Initiation – mRNA and first tRNA bind to ribosome.
Elongation – mRNA is “scanned” by ribosome as the correct
tRNAs react in the correct order.
Termination – The end of the mRNA (a stop codon) is
encountered and translation stops. The newly constructed
protein is released from the ribosome.
Translation Animation
•Amino Acids are connected to each
other using a special kind of covalent
bond called the peptide bond!
•Watch the animation to show how it
happens.
What Happens to the
New Polypeptides?
„
The protein must fold into the correct 3-D shape
‰
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Remember…a denatured protein is an inactive protein!
Some just enter the cytoplasm
Many enter the endoplasmic reticulum and move
through the trafficking system where they are
modified and delivered to distant locations.
Frameshift Mutations
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Insertion
‰
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Extra base added into gene region
Deletion
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Base removed from gene region
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Both shift the reading frame
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Result in altered amino acid sequence
Frameshift Mutation
mRNA
parental DNA
arginine
glycine
tyrosine
tryptophan asparagine amino acids
altered mRNA
arginine
glycine
leucine
leucine
glutamate
DNA with
base insertion
altered aminoacid sequence
Cloning in our Future
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Making a genetically identical copy of an
individual
Numerous species have now been cloned
Mice, pigs, cattle, cats, etc.
Most cloning attempts are still unsuccessful
Many clones have defects.
Would you like to clone your dog? Click here
Cloning in our Future
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How about transgenic organisms?
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These are also called genetically modified
organisms
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Organisms that contain new genes (recipes) put in
place by scientists
New plants that can resist disease
Should we be eating them?
Example showing “proof of principle”
Glow in the dark jellyfish
The Nucleotide Sequence for GFP
1 atggctagca aaggagaaga acttttcact ggagttgtcc caattcttgt tgaattagat
61 ggtgatgtta atgggcacaa attttctgtc agtggagagg gtgaaggtga tgctacatac
121 ggaaagctta cccttaaatt tatttgcact actggaaaac tacctgttcc atggccaaca
181 cttgtcacta ctttctctta tggtgttcaa tgcttttccc gttatccgga tcatatgaaa
241 cggcatgact ttttcaagag tgccatgccc gaaggttatg tacaggaacg cactatatct
301 ttcaaagatg acgggaacta caagacgcgt gctgaagtca agtttgaagg tgataccctt
361 gttaatcgta tcgagttaaa aggtattgat tttaaagaag atggaaacat tctcggacac
421 aaactcgagt acaactataa ctcacacaat gtatacatca cggcagacaa acaaaagaat
481 ggaatcaaag ctaacttcaa aattcgccac aacattgaag atggatccgt tcaactagca
541 gaccattatc aacaaaatac tccaattggc gatggccctg tccttttacc agacaaccat
601 tacctgtcga cacaatctgc cctttcgaaa gatcccaacg aaaagcgtga ccacatggtc
661 cttcttgagt ttgtaactgc tgctgggatt acacatggca tggatgagct ctacaaataa
The Amino Acid Sequence for GFP
MASKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGK
PVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGNYKT
RAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MVLLEFVTAAGITHGMDELYK
Is This a Good Thing?
Zebra fish – “wild type”
transcription
Overview
mRNA
mature mRNA
transcripts
translation
rRNA
ribosomal
subunits
tRNA
mature
tRNA
Animation links
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DNA Structure
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DNA replication
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http://www.johnkyrk.com/DNAreplication.html
Transcription
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http://www.johnkyrk.com/DNAanatomy.html
http://www.johnkyrk.com/DNAtranscription.html
Translation
http://www.johnkyrk.com/DNAtranslation.html