CHAPTER 8 OUTLINE for
STRUCTURE, SYNTHESIS, & REGULATION of DNA, RNA, & PROTEINS
I. Nucleic Acids
A. Types
1. Deoxyribonucleic Acid - DNA
2. Ribonucleic Acid - RNA
RNA
DNA
Single helix
Double helix
5C sugar-ribose
5C sugar-deoxyribose
Bases:
Bases:
Adenine
Adenine
Guanine
Guanine
Cytosine
Cytosine
Uracil
Thymine
B. Historical development of information
1. Johann Miescher (1844-1895) Swiss physician
a. Discovered DNA in 1868 in pus from war bandages and in fish sperm.
b. Called it ‘nuclein’.
2. Robert Fuelgen (1884-1955) German
a. 1914 developed a stain for nuclein
b. “Fuelgen” stain--stains chromatin red
3. Fredrick Griffith (1879-1941) British
a. 1928--worked with streptococcus pneumonia bacteria
b. Hypothesized that DNA was pathogenic
c. Experiment
1. Live Bact R given to mice-- mice lived
1
2. Live Bact S given to mice-- mice died
3. Dead Bact S give to mice--mice lived
4. Live Bact R +dead Bact S--mice died
5. Conclusion: R bacteria got DNA from S bacteria genes?
Transformation?
4. Oswald Avery-(1877-1955) Canadian, MD
a. 1944 used enzymes to destroy protein and then DNA in bacteria.
b. Ist evidence that DNA could transmit disease to rodents, not protein.
c. Concluded that hereditary material was probably made of DNA and not
protein
5. Alfred Hershey (1908-1997) American
Martha Chase (1927-2003)
”
a. Confirmed Avery’s conclusion in 1952
b. Now had radioactive isotopes to work with. Used 32P & 35S
c. Worked with bacteriophage virus T2
1. Labeled the protein coat with 35S
2. Labeled the DNA with 32P.
3. At the end of the experiment, bacteria had 32P in their DNA.
4. Proved that the genes were made of DNA, not protein
6. Erwin Chargaff-(1905-2002) Austrian
a. 1949 worked on DNA composition in USA
b. Concluded that A always equaled T and G = C
c. Also concluded that different species had different numbers of
nucleotides, because they have different amounts of nucleotides
d. Results became known as Chargaff’s Rules
7. Maurice Wilkins (1916-2004) New Zealand
Rosalind Franklin (1920-1958) British
a. 1951--worked in King’s Laboratory in London at Univ of London
b. Both worked on DNA structure in separate labs.
c. Did not get along
d. Franklin made the best X-ray diffraction of crystalline DNA, and worked
out the dimensions of the molecule.
8. James Watson (1928- )
Francis Crick (1916-2004)
a. James Watson--23 year old Ph.D., American
b. Francis Crick--31 year old completing Ph.D., English
c. Became friends at Cavendish Laboratories at Univ. of Cambridge,
England
2
d. Built model of DNA in 1953
e. Used info. of Franklin and her pictures
f. Watson, Crick, and Wilkins awarded Nobel Prize in 1962
g. “Rosie” left England, died of cancer in 1958 at age 37.
The Development of the Watson & Crick DNA Model
1st. problem: Attach the DNA strands with the correct orientation. They oriented the 2 DNA
strands, 5’3’ to 5’3’, but the strands would not bond correctly.
So, Watson & Crick tried this, 5’3’ + 3’5’, & the nucleotides could bond. Actually turned their
paper molecules upside down on the 3’5’ side.
3
2nd Problem: how do the nucleotides bond to each other? Watson & Crick oriented the
nucleotides, Purine to Purine, but the molecule was greater than 2 nanometers:
4
Watson & Crick then oriented the nucleotides this way, Pyrimidine to Pyrimidine, but the width
of the DNA molecule was too narrow.
**************************************************************************
• Chargaff had already determined from DNA samples that
• the number of G purines always equaled the number of C pyrimidines
 AND
• the number of A purines always equaled the number of T pyrimidines.
**************************************************************************
So, Watson & Crick concluded that this was the possible nucleotide bonding in the DNA
molecule.
•
•
Watson & Crick published their paper for the DNA model in 1953.
Watson, Crick, & Wilkins received the Nobel Prize in Physiology or Medicine in 1962 for
the structure of the DNA model. (Rosalind Franklin had already died of cancer.)
5
8. Human Genome Project--decoding human DNA
a. James Watson (DNA model)
1. National Institute of Health-in charge; appointed Watson as
director
2. Headed project in 1988
3. Funded by several countries which participated in the research
4. Watson later resigned due to politics
5. Decoded Watson's DNA in 2004
b. Project completed early 2000 and decoded Watson’s DNA for him in
2004
c. Craig Venter
1. Worked for Celera , Inc.
2. Used computers to speed gene sequencing
3. This was a private company
4. Was NOT a part of the Human Genome
d. Francis Collins
1. NIH 2nd director of government Genome project
2. Rivals with Venter; much animosity
3. Negotiated--jointly announced on June 26, 2000 that the
Human DNA was sequenced.
Genomic studies have advanced rapidly. Many organisms have had their genomes (complete
DNA nucleotide sequence) completed. Used to study disease, genetics, evolution.
III. Structure of DNA
A. Elements--C, H, O, N, P
B. Monomers are called nucleotides
1. Each is made of 3 Subunits
a. Phosphate group
b. Pentose sugar
c. Organic, nitrogenous base
2. Generalized structure of a nucleotide
6
--C. Types of nucleotides--named for the base
1. Pyrimidines--small, 1 carbon ring
a. Cytosine
b. Thymine
2. Purines-large--2 carbon rings
a. Guanine
b. Adenine
D. DNA Structure
1. Double helix
2. Opposite polarity
3. 5’3’ strand and the 3’5’ strand (name comes from the carbon atom numbers in
the sugar)
4. Held together with H bonds bet. the bases.
5. Bases always bond, A-T & G-C.
6. Measurements: 2nm wide, .34nm between pairs, 3.4nm between turns of
helix (10 pairs)
7
E. DNA Replication--Research
1. Watson & Crick
a. Proposed in 1953 that each strand of DNA could act as a template or (pattern)
for replication
b. The H bonds were broken between bases and the molecule “unzipped.”
c. No evidence, just a hypothesis.
2. Arthur Kornberg (1918-2007) USA
a. Used E. coli bacteria in 1957
b. Extracted DNA Polymerase I, an enzyme that catalyzes DNA formation.
c. Added a DNA strand, DNA Polymerase I, and nucleotides with C14.
d. At the end of the experiment, had a radioactive partner for the DNA strand.
e. 1959 Nobel Prize awarded to Kornberg for the discovery of DNA Polymerase I.
8
3. Matthew Meselson (1930-) and Franklin Stahl (1929-)
a. 1958- final conclusive evidence that showed DNA replication was
Semiconservative.
b. Semiconservative – one strand of DNA is made of original nucleotides and the
complimentary strand of DNA is made of new nucleotides.
c. Grew many generations of bacteria in a medium that contained only
radioactive N15. Eventually, all of the N in the DNA of the bacteria
contained N15 or “heavy” nitrogen.
d. Took the N15 and then grew them in “light” radioactive N or N14.
e. In one generation, the DNA was ½ N15 and ½ N14.
f. ½ of the original N15 nucleotides were retained.
4. Semiconservative DNA Replication diagram
Before
During
C-G
T-A
A-T
T-A
T-A
C-G
G-C
T-A
C-G
Original Strands
C-G
T-A
A-T
T-A T-A
T-A T-A
C-G C-G
G-C
G-C
T-A
T-A
C-G
C-G
Orig New
New Orig
After
C-G
T-A
A-T
T-A
T-A
C-G
G-C
T-A
C-G
Orig New
C-G
T-A
A-T
T-A
T-A
C-G
G-C
T-A
C-G
New Orig
5. Steps of Semiconservative Replication
a. Base pairs are complimentary.
Polymerization only in the 5’-3’ direction. Nucleotides added only to the
3’ end.
b. Enzymes-- Helicases unwind the double helix, creating a “fork”.
c. Single strand binding proteins stabilize the unwound molecule.
d. Enzyme--DNA Polymerase III add nucleotides to the leading strand-5’3’
continuously.
e. Enzyme--Primase synthesizes a short RNA primer on the lagging strand 3’-5’
f. Enzyme--DNA Polymerase I adds nucleotides in short sections called Okazaki
fragments.
g. Enzyme--Ligase puts fragments together.
9
F
h. Energy comes from phosphates on free nucleotide.
i. Enzymes proof read--1 mistake / 100,000,000 nucleotides.
“A mistake that survives is a mutation. It usually changes gene. Can cause
disease, improvement, or evolution.”
6. Occurs in Interphase, just before prophase in Mitosis.
7. Repair during replication is controlled by DNA polymerase which can proofread the
copy and cut out errors and substitute the correct base.
8. Mutations can be produced by exposure to radiation, chemicals, and heat.
9. The mutations most likely to survive are spot mutations which involve the
substitution of only one base.
Ex.: cytosine can lose its amino group and be converted to thymine and be
mismatched with G.
HYPP in quarter horses was due to a spot mutation in the quarter horse halter
stallion, Impressive
10. Most mutations can be repaired unless they occur right when the cell is dividing.
11. 50 different enzymes have been identified as having the ability to locate and correct
errors.
G. Organelle DNA – Theory of Endosymbiosis
1. Found in mitochondria and chloroplasts
2. Circular DNA like a prokaryote
3. Very short segment.
4. Attached to the internal membrane of the organelle
5. Allows mitochondria and chloroplasts to replicate.
6. Suggestion: did these organelles once live an independent life?
7. All organelle DNA is contributed by the mother because the egg donates the
cytoplasm for the zygote
8. Therefore, all mitochondrial DNA is maternal DNA.
9. Mitochondria DNA used in evolution studies and forensics
7/2013 announcement: the grave of Richard III has been uncovered in a parking lot demolition
in Leicester, England. Preparation of the body and evidence of the symptoms the king suffered
from (crooked spine) led archeologists to hypothesize that the body of the medieval king had
been uncovered. Mitochondrial DNA from the remains was compared to mitochondrial DNA of
a living female descendant of Richard III in Canada. A match of the mitochondrial DNA
confirmed the identity of the remains as King Richard III.
10