The Chemistry of Inheritance
DNA Structure and
Synthesis
A walk through time
What are the instructions that make a plant? Or a mouse? Or a human?
1860s
Mendel performs experiments and describes the inheritance of
traits in the garden pea.
1870s
Scientists observed the nuclei of eggs and sperm fusing during
fertilization.
1900s
Scientists observed that the number of chromosomes is
constant within a species but varies between species.
Ex. Humans have 46 chromosomes; fruit flies have 8
So, do the chromosomes, which are found in the nucleus, contain the
information that determines inheritance?
Fusion of cytology and genetics
Walter Sutton
Studying grasshopper cells
noticed that chromosomes “come in pairs”
pairs split during meiosis
(see a picture of dividing plant cells)
1902: Chromosomes obey Mendel’s genetic rules
1903: Chromosomes segregate independently
during meiosis, obeying Mendel’s law of
independent assortment
DNA or Proteins: The genetic material?
1920s
Chromosomes contain proteins and DNA.
Most cells of a species contain a constant amount of
DNA, but the amount and kind of proteins vary.
But, is DNA structurally diverse enough to provide the
genetic instructions for an entire organism? Protein
structure seems more varied and therefore more likely to
be able to contain all of the necessary instructions for
heredity…right?
Slimy and Extremely Dangerous
Streptococcus pneumoniae
Encapsulated, “slimy” strains can cause pneumonia (S or
“smooth” strains). Unencapsulated strains do not cause
disease (R or “rough” strains).
Capsule composed of complex carbohydrate.
“R” and “S” strains breed true.
Mice injected with “R” strain or dead “S” strain alone do not get
sick. Mice injected with live “R” strain and dead “S” strain do
get sick, and only “S” strain cells are isolated from the blood of
the mice.
What is converting the “R” cells to “S” cells?
Avery, MacLeod and McCarty
Avery, C.M. et al. 1944. J. Exp. Med. 79, 137.
Culture of
S-strain
bacteria
Kill cells by
heating
Show that the
killed cells do not
grow.
Add killed cells
to culture of Rstrain bacterial
Mostly R
colonies grow;
but a few truebreeding S
colonies appear
Avery, MacLeod and McCarty
Avery, C.M. et al. 1944. J. Exp. Med. 79, 137.
Add protease to
the dead Sstrain cells.
Mix the protease-treated
dead S-strain bacteria
with the R-strain culture.
When plated, you
see mostly Rstrain colonies
and a few S-strain
colonies.
Conclusion: Transforming activity is not protein.
Avery, MacLeod and McCarty
Avery, C.M. et al. 1944. J. Exp. Med. 79, 137.
Add RNase to
the dead Sstrain cells.
Mix the RNase-treated
dead S-strain bacteria
with the R-strain culture.
When plated, you
see mostly Rstrain colonies
and a few S-strain
colonies.
Conclusion: Transforming activity is not RNA.
Avery, MacLeod and McCarty
Avery, C.M. et al. 1944. J. Exp. Med. 79, 137.
Add DNase to
the dead Sstrain cells.
Mix the DNase-treated
dead S-strain bacteria
with the R-strain culture.
When plated, you
see only R-strain
colonies.
Conclusion: Transforming activity is probably DNA.
Hershey Chase Experiment
Phage with labeled DNA direct the production of more phage
that also contain labeled DNA.
Phage with labeled protein do not direct production of more phage with labeled protein.
Chemical Composition of DNA
• DNA is a polymer of nucleotides. Nucleotides consist of:
• deoxyribose (5-carbon) sugar
• nitrogenous base
• phosphate group(s)
Nucleoside
There are four nitrogenous bases used to make the four types of
nucleotides found in a DNA molecule:
Adenine, Thymine, Cytosine and Guanine.
The nucleotides are linked to form a chain. DNA inside cells consists of
two complementary, intertwined chains (double helix).
View a schematic of DNA structure.
Chemical Composition of DNA
• There are four nitrogenous bases that are found in DNA:
adenine, thymine, guanine and cytosine.
• Adenine and thymine are purine bases (2-ring structure)
• Cytosine and guanine are pyrimidines (single-ring
structure)
Chemical Composition of DNA
• The phosphodiester bond of the nucleotide chain is
formed between the phosphate attached to the 5´ carbon
of one sugar and the 3´ carbon of the next.
View the structure of a phosphodiester bond.
• The 5´ end of the strand bears a phosphate group; the 3´
end bears a hydroxyl (OH) group and is considered
“polar”.
• The two strands of DNA in a helical molecule are
antiparallel to each other.
This figure illustrates the polarity of the DNA strands.
Chemical Composition of DNA
• Chargaff’s rules
Base composition varies among species.
Base composition is constant for all cells of an
organism and within a species.
The amount of adenine equals the amount of
thymine.
The amount of cytosine equals the amount of
guanine.
The amount of purine bases equals the amount of
pyrimidine bases.
Structure of the DNA Molecule
• Within cells the standard structure of DNA is the B form.
• The B form structure consists of two antiparallel
polynucleotide chains twisted around one another to form
a double helix.
• The nitrogenous bases form the “rungs” in the center of
the helix, with adenine forming hydrogen bonds with
thymine and guanosine forming hydrogen bonds with
cytosine.
• The helix is right-handed, and each chain makes one
complete turn every 34 angstroms.
DNA Replication: Meselson and Stahl
• Studied DNA replication in E. coli.
• Cells were grown in “heavy” medium in which DNA
molecules would incorporate 15N (a heavy isotope of
nitrogen).
• They were transferred to “light” medium, which contained
only 14N (“light”) nitrogen.
• DNA samples were taken at various time points subjected
to centrifugation on a density gradient.
• Molecules containing only 15N would run lower on the
gradient than molecules containing 15N + 14N or 14N alone.
Evidence for Semi-Conservative Replication
• At time 0 all of the DNA banded together as “heavy” DNA
• One generation after switching the bacteria to the light
medium, half-labeled or “hybrid” molecules were observed
in the density gradient.
• The gradual appearance of half-labeled and unlabeled
molecules on the density gradient led to these
conclusions:
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The nitrogen of a DNA molecule is divided equally between two
subunits…following rpelication each daughter molecule has
received one parental subunit…the replicative act results in a
molecular doubling. (Meselson and Stahl (1958) Proc. Natl. Acad.
Sci. USA 44, 671–82.)
A schematic of the experiment is available here.
DNA Polymerase
• DNA polymerase catalyzes the chemical reaction that
joins the 5´ phosphate with the 3´ OH to form the
phosphodiester bond and, hence, the polynucleotide
chain.
• DNA polymerases require:



The 5´ triphosphates of the four deoxynucleosides (dATP, dGTP,
dCTP, dTTP)
A pre-existing single strand of DNA (template)
A primer—short segment of RNA (in cells) that has a free 3´ OH
group
Polymerase Proofreading
• Not only does DNA polymerase catalyze the addition of
new nucleotides to a growing molecule, it can also
remove nucleotides.
• DNA pol III has a 3´ to 5´ exonuclease activity that allows
it to remove incorrectly incorporated nucleotides from the
end of the growing molecule. This is the proofreading or
editing function of DNA polymerase.
Replication
• Several events must happen before DNA polymerase can
catalyze the formation of a new DNA molecule based on
an existing template.



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The double helix must unwind to expose the two template strands.
This is usually accomplished by the action of another enzyme,
helicase.
Once the helix is unwound it must be “stabilized” in this open state.
This is accomplished by single-stranded binding protein.
Before DNA polymerase can extend a strand, it must have a short
primer. The primer is synthesized by an RNA polymerase called
primase.
The area of the helix that is unwound for replication is the
replication fork.
Leading and Lagging Strand Synthesis
• DNA polymerase can only add to a free 3´ OH.
• The two template strands are antiparallel.
• Therefore, only one of the strands can provide a template
for continuous 5´ to 3´ new strand synthesis. The strand
synthesized from this template is called the leading
strand. It is synthesized in one piece in the direction of the
replication fork.
• The daughter molecule that is synthesized from the other
half of the helix is the lagging strand. It is synthesized in
short fragments, called Okazaki fragments. These
fragments are joined to form the complete daughter
molecule.