Download DNA and Replication (Chapter 16)

11/24/2015
Chapter 16
P. 305 - 324
16.1 Dna Is The Genetic Material
Important Scientists in the Discovery of DNA
 T.H Morgan’s group:
 Frederick Griffith
 showed that genes are located along chromosomes.
 Two chemical components of chromosomes are DNA
and protein.
 Oswald Avery
 Alfred Hershey and Martha Chase
 Little was known about nucleic acids.
 Rosalind Franklin
 Role of DNA in heredity was first worked out by
studying bacteria and the viruses that infect them.
 Francis Crick and James Watson
Frederick Griffith
Frederick Griffith




 This newly acquired trait
Discovery of role in 1928
Vaccine against pneumonia (mice)
Frederick Griffith studied Streptococcus pneumoniae
Two stains of the bacterium
 Pathogenic
 Non pathogenic
 Heated the pathogenic and killed the bacteria.
 Mixed the cell remains with living bacteria of the
nonpathogenic and found some cells were then
pathogenic
was inherited by all the
descendants of the
transformed bacteria.
 Called the phenomenon
transformation:
 a change in genotype and
phenotype due to the
assimilation of external
DNA by a cell.
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Oswald Avery
Oswald Avery
 Identity of transforming substance
 Three main candidates
 DNA
 RNA
 Protein
 Avery broke open the heat-killed bacteria and
extracted the cellular contents
 Special treatments to inactivate each of the three
molecules
 Tested each for its ability to transform live
nonpathogenic bacteria.
 DNA was left active – transformation occurred
 Transforming agent was then announced as DNA
 Studied viruses for more information
 Bacteriophages (phages): bacteria-eaters
 A virus is composed of DNA(or RNA) enclosed by
a protective coat.
Fig. 16-4-3
EXPERIMENT
Hershey and Chase
 Devised an experiment
showing that only one of
the two components
enters the E.coli cell.
 Specifically looked at T2
 T2 invades Escherichia
coli bacteria
 Radioactive isotope of
sulfur to tag protein, and
phosphorus to tag DNA.
Rosalind Franklin
Phage
Empty
Radioactive protein
shell
protein
Radioactivity
(phage
protein)
in liquid
Bacterial cell
Batch 1:
radioactive
sulfur (35S)
DNA
Phage
DNA
Centrifuge
Pellet (bacterial
cells and contents)
Radioactive
DNA
Batch 2:
radioactive
phosphorus (32P)
Centrifuge
Pellet
Radioactivity
(phage DNA)
in pellet
Francis Crick and James Watson
 Used X-Ray crystallography

to find out structure of
DNA molecules
 X near center shows DNA
twists around
 Angle of the X suggests two
strands and the
nitrogenous bases are near
the center of the molecule
 Shows diameter of the
double helix

Built three-dimensional models of DNA
Used Rosalind Franklin’s x-ray pictures of DNA to
assist in the model
 The Double Helix
 Width suggested that it was made up of two
strands.
 Began to build models that would conform to the
X-ray measurements and the chemistry of DNA.
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Watson and Crick- Double Helix
Chargaff’s Rule
 Composed of two complementary
strands of DNA wrapped around each
other
 Studied percentages of nitrogenous bases.
 Almost equal %’s
 Adenine bonds to Thymine – Guanine bonds to
 Uniform diameter
 Hydrogen bonds held the two strands
together
Cytosine = support
 Nitrogenous Bases make up DNA molecules
 The two types are:
 Purines
 Two hydrogen bonds between A and T
 Three hydrogen bonds between C and
G.

Two rings in the structure
 Pyrimidines
 One ring in the structure
Fig. 16-5
Sugar–phosphate
backbone
5 end
Nitrogenous
bases
Chargaff’s Rule
Thymine (T)
Adenine (A)
Cytosine (C)
DNA nucleotide
Phosphate
Sugar (deoxyribose)
3 end
Guanine (G)
Base Pairing
 Watson and Crick stated their hypothesis
 Pair of templates, each of which is complementary to the
other.
 Prior to duplication, the hydrogen bonds are broken
 The two chains unwind and separate
 Each chain acts as a template
 Eventually, two pairs of chains will result.
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DNA Replication
 In DNA replication, the parent molecule unwinds, and
two new daughter strands are built based on basepairing rules
 When a cell copies a DNA molecule, each strand serves as a
template for ordering nucleotides into a new, complementary
strand.
 Nucleotides line up along the template strand and are linked
 Where there was one double-stranded DNA molecule at the
beginning, there are then two at the end.
 The copying mechanism is analogous to using a photographic
negative to make a positive
Watson and Crick’s Hypothesis
 Figure 16.9
A
T
A
T
A
T
A
T
C
G
C
G
C
G
C
G
A
T
A
T
A
T
A
T
A
T
A
T
A
T
A
T
G
C
G
C
G
C
G
C
(a) Parent molecule
(b) Separation of
strands
Fig. 16-10
Parent cell
Replication Models
(c) “Daughter” DNA molecules,
each consisting of one
parental strand and one
new strand
First
replication
Second
replication
(a) Conservative
model
 Remained untested for many years
 Difficult to perform
 Watson and Crick predicted the semiconservative
model
(b) Semiconservative model
 Each daughter molecule will have one old strand
(derived or “conserved” from the parent molecule) and
one newly made strand
 There are two others:
 Conservative
 Dispersive
(c) Dispersive
model
DNA Replication Models
Semiconservative Model
 Conservative
 The two parental strands reassociate after acting as
templates for new strands.
 Semiconservative
 The two strands of the parental molecule separate, and
each functions as a template for synthesis of a new,
complementary strand.
 1950
 Dispersive
 Each strand of both daughter molecules contains a
mixture of old and newly synthesized DNA.
 Matthew Meselson and Franklin Stahl devised a clever
experiment that supported the semiconservative
model.
 Widely acknowledged among biologists to be a classic
example of elegant experimental design.
 Figure 16.11 shows the experiment performed by
Meselson and Stahl.
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DNA and Replication in Prokaryotes
Prokaryotes
 Prokaryotes:
 ring of chromosome
 holds nearly all of the cell’s
genetic material
 DNA replication begins
at a single point and
continues to replicate
whole circular strand
 Replication goes in both
directions around the
DNA (begins with
replication fork)
Eukaryotes
Eukaryotic DNA Replication
Eukaryotic DNA Replication
 The replication of a DNA molecule begins at
special sites called origins of replication
 Hydrogen bonds between base pairs breaks
 Begins in hundreds of locations along the
chromosome
 Begins when the DNA molecule “unzips” creating:
 Replication fork
 Replication “bubble”
 Helicases – enzymes that untwist the double helix
at the replication forks.
 Single-strand binding proteins bind to the
unpaired DNA strands, stabilizing them.
 Topoisomerase – relieves pressure of DNA ahead
of replication fork
 RNA Primer – RNA chain
 Primase – enzyme that synthesizes the primer
Synthesizing a New DNA Strand
 Helicase will start to unwind the DNA strand.
 Topoisomerase will hold the strands together and prevent
breaking
 Single-stranded binding proteins will stabilize the DNA
 DNA polymerase: catalyze the synthesis of new
DNA by adding nucleotides to a preexisting chain.
 Most DNA polymerases require a primer and a DNA
template strand.
 DNA polymerase III adds a DNA nucleotide to the
RNA primer and then continues adding DNA
nucleotides complementary to the parent DNA
template strand.
strands
 The Primase will start to form the RNA chain
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Antiparallel Elongation
Antiparallel Elongation
 The two strands of DNA in a double helix are
 Leading Strand –only 1 primer needed, moves toward
antiparallel (0riented in opposite directions).
 DNA polymerases can only add nucleotides to the free
3’ end of a primer or growing strand.
 Lagging Strand – many primers needed, moves away
 NEVER THE 5’
 A new strand can only elongate in the 5’-3’ direction
 ALWAYS
 Read in the 3’ – 5’ direction
 Created in 5’ – 3’ direction
the replication fork
from the replication fork
 Okazaki Fragments – on lagging strand, short
segment of DNA synthesized away from the
replication fork
 DNA ligase – enzyme, joins the sugar-phosphate
backbones of all the Okazaki fragments into a
continuous DNA strand
The DNA Replication Complex
 By interacting with other proteins at the fork,
primase acts as a molecular brake, slowing
progress of the replication fork.
 The DNA replication complex does not move
along the DNA
 The DNA moves through the complex
Proofreading and Repairing DNA
 During DNA replication, DNA polymerases proofread
each nucleotide against its template as soon as it is
added to the growing strand.
 The polymerase removes the incorrectly paired
nucleotide and resumes synthesis.
 Mismatched nucleotides sometimes are missed.
 Can also arise after replication
 Mismatched repair – enzymes remove and replace
incorrectly paired nucleotides that have resulted from
replication errors.
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Proofreading and Repairing DNA
Replicating the Ends of DNA
Telomeres
 Most cellular systems that repair incorrectly paired
 Found at the ends of each chromosome and contain no
nucleotides use a mechanism that takes advantage of
the base-paired structure of DNA.
 Nuclease – DNA-cutting enzyme.
 Telomerase lengthens telomeres in gametes
 Cuts out the segment of the strand containing the
damaged segment.
 Enzymes involved in filling gaps:
 DNA polymerase and DNA ligase
 Nucleotide excision repair – repair system, Figure
genes (protective cap)
 Adds DNA bases at the 5’ end
 The shortening of telomeres might protect cells from
cancerous growth by limiting the number of cell
divisions
16.18
Important Enzymes to Remember
 Helicase, single-strand binding protein, topoisomerase
 Primase
 Synthesis of RNA primer
 DNA polymerase III (DNA pol III)
 Add new bases to DNA strand
 DNA polymerase I (DNA pol I)
 Removes and replaces RNA primer from 5’ end
 DNA ligase
 Links Okazaki fragments and replaces RNA primer from 3’ end
Vocab
Chromosome Structures
 Nucleoid –A dense region of DNA in a prokaryotic
 Bacteria:
 one double-stranded, circular DNA molecule that is
associated with a small amount of protein.
 Prokaryotes:
 Ring of chromosomes
 Holds nearly all the cell’s genetic material
 Eukaryotes:
 DNA in chromosomes
 Found in nucleus
cell
 Chromatin –complex of DNA and proteins that
makes up a eukaryotic chromosome
 Heterochromatin – Eukaryotic chromatin that
remains highly compacted during interphase and is
generally not transcribed.
 Euchromatin – The less condensed form of
eukaryotic chromatin that is available for
transcription.
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Fig. 16-21a
Chromatin Packing
 In the cell, eukaryotic DNA is combined with large
amounts of protein.
 Complex of DNA and protein – chromatin
 Histones - proteins that are responsible for the
first level of DNA packing in chromatin
 Form a tight bond because DNA is negatively
charged and the histones have a positive charge
Nucleosome
(10 nm in diameter)
DNA
double helix
(2 nm in diameter)
H1
Histones
DNA, the double helix
Histones
Histone tail
Nucleosomes, or “beads
on a string” (10-nm fiber)
Fig. 16-21b
Chromatid
(700 nm)
30-nm fiber
Chromosome Organizations
 10-nm fiber
Loops
 30 – nm fiber
Scaffold
 300-nm fiber
300-nm fiber
Replicated
chromosome
(1,400 nm)
30-nm fiber
Looped domains
(300-nm fiber)
Metaphase
chromosome
10 - nm fiber
 DNA winds around histones to form nucleosome beads
 Nucleosomes are strung together
 The string between the beads is called linker DNA
 Nucleosome consists of DNA wound twice around a
30-nm fiber
 10-nm coils
 Forms a chromatin
fiber
 30 nm thick
 Interactions between
nucleosomes cause
the thin fiber to coil
or fold into this
thicker fiber
protein core composed of two molecules each.
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300-nm Fiber
 30 nm fiber forms loops
called looped domains
attached to a
chromosome scaffold
made of proteins
 Scaffold is rich in one
type of topoisomerase.
Heterochromatin and Euchromatin
 Heterochromatin
 During interphase, a few regions of chromatin are highly
condensed into heterochromatin

 Euchromatin
 Most chromatin is loosely packed in the nucleus during
interphase

Questions
Dense packing of the heterochromatin makes it difficult for
the cell to express genetic information coded in these regions
Condenses prior to mitosis
DNA ligase, DNA polymerase, Helicase, Primase,
Telomerase
Single-strand binding proteins, Topoisomerase,
Nuclease
_______ removes section of DNA that is damaged
_______ proofreads and repairs damaged/mismatched
DNA; base pairing
3. _______ synthesis of RNA primer
4. _______ Links Okazaki fragments; replaces RNA primer
from 3’ end (in both leading and lagging strand).
5. _______ relieves pressure of DNA ahead of replication fork
6. _______ attach to separated DNA strands to ensure they
stay separated
7. _______ breaks hydrogen bonds between DNA strands
8. _______ lengthens telomeres in gametes
1.
2.
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