INTRODUCTION TO
DNA Replication
-------------------------------------------------Chapter 13 covers a descriptive explanation of Deoxyribose nucleic Acid (DNA) and the replication of
DNA. Chapter 14 covers the transcription of the genetic information embedded in DNA to a special type
of Ribose Nucleic Acid (m RNA) for later translation into proteins which express heritable characteristics
we are all familiar with.
Reference:
Chapter 13 & 14 of Starr & Tagart. 10th Ed.
1. http://www.pathology.washington.edu/Cytogallery
This site defines and describes a Genetic material
2. http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookDNAMOLGEN.html
3.. http://www.accessexcellence.com/AB/GG/protein_synthesis.html
This site provides narrative and pictures of protein synthesis process
4. http://web.indstate.edu/thcme/mwking/rna.html
An Excellent site for explanations for and definitions of the protein synthesis process
5. http://photoscience.la.asu.edu/photosyn/courses/BIO_343/lecture/DNA-RNA.html
Out standing drawing of the sugars of DNA and RNA (a comparison)
6. http://molbio.info.nih.gov/molbio/gcode.html
This last web site is the genetic code which you can also find in text book
Learning Objectives:
1. Describe the biochemical, structural, geometric aspects of DNA.
a. How does hydrogen bonding play a very big role in determining the pairing of the 4 bases
composing DNA
2. Explain how DNA replicates
a. Know why it is said DNA replication is semi-conservative in nature.
3. Explain “What are genes” in a biochemical sense.
4. Describe how copied genetic information from DNA, which never leaves the nucleus, is able
to find its way into the cytoplasm for the construction a polypeptide units.
5. Know the various forms of RNA in the transcription, translation, elongation & termination
process of a polypeptide chain.
6. Be able to decode and demonstrate the use the “GENETIC CODE” for the synthesis of
amino acids from given codon sequences.
7. Fully understand the differences between DNA replication and DNA transcription/translation
processes.
8. Understand the consequences of mutations, deletions, or frame shift reading errors that arise
9. Know the vocabulary associated with both chapters.
---------Discussion Question
1. The Central Dogma states that DNA codes for polypeptides by opening a section of the double helix
for transcription of the genetic information to a single strand of m RNA. The m RNA is then exported
from the nucleosome into the cytoplasm for processing, translation, into peptides. Discuss how the
Human Immnodefiency Virus (HIV), an RNA virus, is able to reverse this Central Dogma concept by
incorporating a piece of its genetic material into a host’s genomic structure (DNA). How the possibilities
this process being used in future gene therapies procedures.
Chapter 13 Notes
I. DNA Structure
A. Components of DNA
1. DNA is composed of 4 kinds of nucleotides, each of which consists
of:
a. A five-carbon sugar - deoxyribose
b. A phosphate group
c. One of four bases:
Adenine (A)
Guanine (G)
Thymine (T)
Cytosine ©)
2. There are 2 classes nucleotides that make DNA
a. Pyrimidine:
single-ring thymine and cytosine
b. Purines:
double ring adenine and guanine
3. DNA composed of 4 kinds of nucleotide bases pairing in an
A~T
and
G~C
fashion through the process of hydrogen bonding
A ~ T forms two hydrogen bonds
G ~ C forms three hydrogen bonds
a. These base pairs differ in relative amounts from species to species
4. DNA exist as long, thin molecule of uniform diameter
a. The structure is highly repetitive
b. DNA is helical
B. Patterns of base Pairing
Watson and Crick model of DNA
1. Single ring 'T' hydrogen bonded with double ringed 'A' and single - ringed 'C' with
double-ringed 'G'
Examine the hydrogen bond linkage. This is a key factor here!
2. The backbone was made of chains of sugar-phosphate linkages
3. The molecule was double stranded and looked like a ladder with a twist to form a
double helix
4. This base pairing is constant for all living species however the base pairing is
different between species
II. DNA Replication and Repair
A. Making of Nucleotide Strands
1. Two strands of DNA unwind and expose their bases
2. Unattached nucleotides pair with exposed bases
3. Replication results in DNA molecules that consist of one "old" strand and one
"new" strand; Note this is considered a 'semi-conservative replication' method
B. Enzymes for Replication
1. One kind of enzyme unwinds the 2 nucleotide strands
2. DNA polymerase attach free nucleotides to the growing strand
3. DNA ligase seal new short stretches of nucleotides into one continuous strand
C. DNA polymerase, DNA ligase, and other enzymes engage in DNA repair
D. DNA polymerase "proof read" the new bases for mismatched pairs and will replace
mismatches with correct base
III. DNA Organization
A. DNA of eukaryotes is highly organized to prevent tangling
1. Some histones (a type of protein) as spools to wind the DNA into units called
nucleosomes
2. Another histone (H1) stabilizes the arrangement
a. Beaded chain forms looped regions
3. Other proteins serve as structural scaffolds for loops
a. This arrangement may allow for functional "domains"
Chapter 13
DNA Structure and Function
DNA exists in the nucleus as a condensed, compact structure. To prepare DNA for
replication, a series of proteins aid in the unwinding and separation of the
double-stranded DNA molecule. These proteins are required because DNA must be
single-stranded before replication can proceed.
Unwinding protein: Helicases - These proteins bind to the double stranded DNA and
stimulate the separation of the two strands.
Strand separating protein: Single strand binding protein - These proteins bind to the
DNA as a tetramer and stabilize the single-stranded structure that is generated by the
action of the helicases. Replication is 100 times faster when these proteins are
attached to the single-stranded DNA.
Untangling proteins: Gyrase or topoisomerase -This enzyme catalyzes the formation of
negative supercoils that is thought to aid with the unwinding process. In addition to
these proteins, several other enzymes are involved in bacterial DNA replication.
Stress releasing protein
These topoisomerase proteins act by breaking DNA strands thereby
allowing the molecule to rotate around a single strand
Primase: This enzyme is a short stretch of RNA (5 sequence long) which
binds to complementary DNA base pair
1) RNA contains uracil in place of thymine
2) Also has ribose sugar instead of deoxyribose sugar
DNA polymerase: This enzyme catalyze the synthesis of a new DNA strand
RNA polymerase, on the other hand, synthesizes mRNA DNA Polymerase
- DNA Polymerase I (Pol I) was the first enzyme discovered with
polymerase activity, and it is the best characterized enzyme. Although this
was the first enzyme to be discovered that had the required polymerase
activities, it is not the primary enzyme involved with bacterial DNA
replication. That enzyme is DNA Polymerase III (Pol III). Three activities
are associated with DNA polymerase I;
* 5' to 3' elongation (polymerase activity)
* 3' to 5' exonuclease (proof-reading activity
* 5' to 3' exonuclease (repair activity)
The second two activities of DNA Pol I are important for replication, but
DNA Polymerase III (Pol III) is the enzyme that performs the 5'-3'
polymerase function.
DNA polymerase III links nucleotides in a 5' to 3' direction
Energy to form the new covalent bonds comes from release of pyrophosphate
from ATP
-DNA strands are formed in an anti-parallel fashion
New DNA synthesis occurs in a 5' to 3' direction
One strand (leading strand) is made in continuous order on the 3' - 5'
template while the other strand (lagging strand) is synthesized in short
segments.
These segments are called OKAZAKI fragments which are usually 100
to 300 nucleotides long in Eukaryotes.
Each fragment is primed with RNA primes
DNA polyermase II removes the primer & replaces it with DNA
DNA Ligase - Nicks occur in the developing molecule because the RNA
primer is removed and synthesis proceeds in a discontinuous manner on
the lagging strand. The final replication product does not have any nicks
because DNA ligase forms a covalent phosphodiester linkage between
3'-hydroxyl and 5'-phosphate groups.
Proof reading the DNA : There is a high degree of accuracy in DNA
replication
In bacteria there can be an error in the order of 1:10,000 base pairs
DNA polymerase I backs up and removes the corrupted nucleotide
sequence in a 5' - 3' direction
The analysis of temperature-sensitive mutants of E. coli has defined
a series of genes and their role in DNA synthesis. The following
table list some of the genes and their role in E. coli DNA replication.
Gene
Function
dnaA,I,P
Initiation
dnaB,C
Helicase at oriC
dnaE,N,Q,X,Z
Subunits of DNA polymerase III
dnaG
Primase
gyrA,B
Subunits of gyrase
lig
Ligase
oriC
Origin of Replication
polA
DNA polymerase I
polB
DNA polymerase II
rep
Helicase
ssb
Single-stranded DNA binding proteins
F:\Biology_Book\Chapter 13\Notes Introductiion\CHAP_13_Notes.wpd
Chapter 13 Study Guide Material
Name:_____________________________
Match the numbers on the illustration below to the letters representing structures and functions.
6.____ A. The enzyme (ligase) links short DNA segments together.
7. ____ B. One parent DNA strand.
8. ____ C. Enzymes unwind DNA double helix.
9. ____ D. Enzymes add short “primer” segments to begin chain assembly
10. ___ E. Enzymes (DNA polymerase) assembles new DNA strands.
11. ___ F. Newly-forming DNA strand.
True-False
If the statement is true, write a T in the blank. If false, correct it by changing the underlined word(s) and
writing the correct word( s) in the blank.
____ 13. DNA is composed of four different types of nucleotides.
____ 14. In the DNA of every species, the amount of adenine present always equals the amount of
thymine, and the amount of cytosine always equals the amount of guanine (A = T and C = G).
____ 15. In a nucleotide, the phosphate group is attached to the nitrogen-containing base which is
attached to the five-carbon sugar.
____ 16. Watson and Crick built their model of DNA in the early 1950S.
____ 17. Guanine pairs with cytosine and adenine pairs with Thymine by fanning hydrogen bonds
between them.
Label-Match
Identify each indicated part of the righthand DNA illustration. Choose from these answers: phosphate group, purine,
pyrimidine, nucleotide, and deoxyribose. Complete the exercise by matching and entering the letter of the proper
structure description in the parentheses following each label.
The following DNA memory devices may be helpful: Use pyrCUT to remember that the single ring pyrimidines are
cytosine, uracil, and thymine; use purAG to remember that the double-ring purines are adenine and guanine;
pyrimidine is a long name for a narrow molecule; purine is a short name for a wide molecule; to recall the number of
hydrogen bonds between DNA bases, remember that AT = 2 and CG = 3.
6.________________________________________ (
)
7. ________________________________________ (
)
________________________________________ (
)
8. ________________________________________ (
)
9. _______________________________________ (
)
10. ______________________________________ (
)
11. _____________________________________ (
)
12. _____________________________________ (
)
A. The primidine is thymine because it has two hydrogen bonds.
B. A five-carbon sugar joined to two phosphate to two phosphate
Groups in the upright portion of the DNA ladder.
C. The purine is guanine because it has three hydrogen bonds.
D. The pyrimidine is cytosine because it has it has three
hydrogen bonds.
E. The purine is adenine because it has two hydrogen bonds.
F. Composed of three small molecules: a phosphate group, fiveCarbon deoxyribose sugar, and a nitrogenous base (in this case, a
Pyrimidine).
G. A chemical group that joins two sugars in the upright portion
of the DNA ladder.
Labeling
Four nucleotides are illustrated below. In the blank, label each nitrogencontaing base correctly as guanine,
thrymine, cytosine, or adenine. In the parentheses following each blank, indicate whether that nucleotide
base is a purine (pu) or a pyrimidine (py).
Label
The term semiconservative replication refers to the fact that each new DNA molecule resulting
from the replication process is "half-old, half-new." In the illustration below, complete the
replication required in the middle of the molecule by adding the required letters representing the
missing nucleotide bases. Recall that ATP energy and the
appropriate enzymes are actually required in order to
complete this process.
T
G
A
C
C
C
old
___
___
___
___
___
___
new
A
C
T
G
G
G
old
___
___
___
___
___
___
new
True-False
_____ 1. The hydrogen bonding of adenine to guanine
is an example of complementary base pairing.
_____ 2. The replication of DNA is considered a
conserving process because the same four
nucleotides are used again and again during
replication.
______ 3. Each parent strand remains intact during
replication, and a new companion strand is
assembled on each of those parent strands.
______ 4. Some of the enzymes associated with DNA
assembly repair errors during the replication process.
Self-Quiz
1. Each DNA strand has a backbone that consists of alternating
a. purines and pyrimidines b. nitrogen-containing bases
c. hydrogen bonds
d. sugar and phosphate molecules
2. In DNA, complementary base pairing occurs between
a. cytosine and uracil
b. adenine and guanine
c. adenine and uracil
d. adenine and thymine
3. Adenine and guanine are
a. double-ringed purines,
b. single-ringed purines
c. double-ringed pyrimidine d. single-ringed pyrimidines
4. Franklin used the technique known as to determine many of the physical characteristic of DNA.
a. transformation
b. transmission electron microscopy
c. density-gradient centrifugation d. X-ray diffraction
5. The significance of Griffith's experiment that used two strains of pneumonia-causing bacteria is that
a. the conserving nature of DNA replication was finally demonstrated
b. it demonstrated that harmless cells had become permanently transformed through a change in the
bacterial hereditary system
c. it established that pure DNA extracted from disease-causing bacteria trans- formed harmless strains
into "pathogenic strains"
d. it demonstrated that radioactively labeled bacteriophages transfer their DNA but not their protein
coats to their host bacteria
6. The significance of the experiments in which 32P and -355 were used is that
a. the semiconservative nature of DNA replication was finally demonstrated
b. it demonstrated that harmless cells had become permanently transformed through a change in the
bacterial hereditary system
c. it established that pure DNA extracted from disease-causing bacteria trans- formed harmless strains
into "killer strains"
d. it demonstrated that radioactively labeled bacteriophages transfer their DNA but not their protein
coats to their host bacteria
7. Franklin's research contribution was essential in
a. establishing the double-stranded nature of DNA
b. establishing the principle of base pairing
c. establishing most of the principal structural features of DNA
d. all of the above
8. When Griffith injected mice with a mixture of dead pathogenic cells-encapsulated 5 cells and living,
unencapsulated R cells of pneumonia bacteria- he discovered that
a the previously harmless strain had permanently inherited the capacity to build protective capsules
b. the dead mice teemed with living pathogenic (R) cells
c. the killer strain R was encased in a protective capsule
d. all of the above
9. A single strand of DNA with the base-pairing sequence C-G-A-T-T-G is compatible only with the
sequence
a. C-G-A-T-T-G b. G-C-T-A-A-G c. T-A-G-C-C-T d. G-C-T-A-A-C
10. Which of the following statements is not true?
a. The DNA molecules of each species show unique differences in the sequence of base pairs along
their lengths.
b. One strand of a DNA molecule runs in the 5'--> 3'direction and the other runs in the 3' -> 5' direction.
c. During cell division, enzymes unwind and separate DNA's two strands. Each strand remains intact
throughout the process and enzymes assemble a new, complementary strand on each parent strand.
d. Enzymes involved in replication also repair base-pairing errors which have appeared in the
nucleotide sequence of DNA.