DNA:BACTERIOPHAGE DNA : VIRUSES

-Hershey and Chase

-It was first demonstrated that
radioactively labeled phage
when purified RNA from
DNA with phosphorus-32 (32P)
tobacco mosaic virus was
and labeled phage protein with
spread on tobacco leaves, the
sulfur-35 ( 35 S).
leaves showed lesions of viral

- This is because DNA has infection.

phosphorus but not sulfur,

- Some groups of viruses are
whereas protein contains
known to use DNA as their
sulfur, but not phosphorus.
hereditary material, such as

-Hershey and Chase let the the T2 bacteriophage in

labeled T2 bacteriophages Hershey and Chase’s

infect the unlabeled bacteria experiment

and inject their genetic

NUCLEIC ACIDS COMPONENTS
material into the cells.
POLYNUCLEOTIDES
-Direct evidence that DNA is
the genetic material in - Macromolecules nucleic acids
eukaryotes comes from that exist as polymers.
recombinant DNA technology.
NUCLEOTIDES
GENE CLONING technique is
- A polynucleotide that
now widely used in current
consists of many monomers
biomedical research and
pharmaceutical production - Considered the building
block of all nucleic acid
molecules.
-These structural units
nucleic acids consist of three
essential components:
 nitrogenous base
 phosphate group
 pentose sugar
NITROGENOUS BASE
- The five-carbon sugar ring
and the content of the
nitrogenous base between
DNA and RNA are slightly
different from each other.
Four different types of
nitrogenous bases are found in
DNA:
 Adenine (A)
 Thymine (T)/RNA: Uracil
 Cytosine (C)
 Guanine (G)
PHOSPHATE GROUP
NUCLEOSIDE - Composed of a
purine or pyrimidine base and
a ribose or deoxyribose sugar.
• Nitrogenous base and the
pentose sugar are linked by
glycosidic bond.
 PURINE BASE: N-9 atom
bonded to the sugar
 PYRIMIDINE: N-1 atom
bonded to the sugar
of NUCLEOTIDE
- a phosphate group attaches
to a nucleoside through a
phosphoester bond ( 5’-
hydroxyl group of the sugar
and a phosphate group)
 Phosphomonoester
bond
 Phosphate group + (1)
sugar =
Phosphomonoester
bond
NUCLEOTIDE FORMATION
NUCLEOSIDE
TRIPHOSPHATES
- Serves as the precursor
molecule during nucleic acid
synthesis within the cell.
- The hydrolysis of ATP to
ADP, releasing one
mol_x0002_ecule of inorganic
phosphate (Pi), is accompanied
by the release of large amount
of energy in the cell.
- Phosphodiester bonds can be
found in both DNA and RNA.
- One end of the
polynucleotide has a free 5’-
phosphate group, so it is
called the 5’-end.
- The other end of the ANGSTROM (Å)
polynucleotide has a free 3’-
hydroxyl group, and it is called unit of length, equal to 10−10
3’-end. metre, or 0.1 nanometre.

- This figure is read as “TCA

THE STRUCTURE OF
DNA MOLECULES
1953, James Watson and
Francis Crick

proposed the theory

that DNA molecules
exist as a double helix

TWO MODELS OF DNA STRUCTURE

The DNA double helix is

presented as a twisted ladder.

A space -filling model of DNA

structure.

Rosalind Franklin and

Maurice Wilkins

The DNA model is based

on the X -ray diffraction.

Erwin Chargaff

Observed DNA composition

studies from the data
collected by Franklin and
Wilkins.
 Chargaff’s Rules 9. The percentage of (G+C) is
not necessarily equal the
1. Two long polynucleotide percentage of (A+T)
chains are coiled around a
central axis, forming a right-
handed double helix. In combination of Chargaff’s
rules with the fact that DNA
2. The two DNA strand are
structure has a 3.4 A˚
antiparallel, that is, their
periodicity.
5’→3’ orientation runs in
opposite directions. Watson and Crick proposed
that DNA must be a double
3. The base of both chains lie
helix with its sugar-phosphate
perpendicular to the axis, and
backbone on the outside and
they are stacked on one
its nitrogenous bases on the
another.
inside.
4. The nitrogenous bases of
BASE PAIR
opposite chains are paired as
the result of the formation of a • The two polynucleotide
hydrogen bond in DNA. chains in the double helix are
held by hydorgen bonding
5. Each complete turn of helix
between nitrogenous bases.
is 34 A˚ long.
For DNA, G bond ONLY with C
6. The double helix has a
; A can bond ONLY with T
diameter of 20 A˚ .

7. The amount of adenine (A)

residues is proportional to the DNA STRUCTURE:
amount of thymine (T) Arrangement of each subunit
residues in DNA. Also, the
amount of guanine (G) The Watson-Crick model places
residues is proportional to the the sugar-phosphate
amount of cytosine (C). backbones on the outside of
the double helix and carries
8. The sum of the purines the negative charges on the
equal to the sum of phosphate group. ANTI-
pyrimidine. PARALLEL • two polynucleotide
chains run in opposite
directions.
NITROGENOUS BASES DNA: Right-handed, clockwise
twist; ~10 base pairs per turn
- inside of the double helix. for average structure:

- flat and perpendicular to the OVERWOUND: more base

axis of the helix. pairs per turn

- stacked above one another UNDERWOUND: fewer base

around the axis like spiral pairs per turn
staircase.

STAIRS: Nitrogenous bases

CURVING SIDES: Sugar-
phosphate backbones of the
two DNA strands
Twisting of two strands to one
another resulting in MINOR
AND MAJOR GROOVES
ALTERNATIVE FORMS OF DNA
“B-DNA”
• A DNA molecule that exists
in a very high relative humidity
environment (92%).
• the double helix is said to
be right handed.

both can be sites at which • B-DNA is the most common

drugs or polypeptides bind to form in vivo and in solution in
DNA. vitro • CONVERSION

 Minor: distance is ~12 • 1 Angstrom = 0.1

Å; Nanometre
 Major: distance ~22 Å

DISTANCE OF:
RIGHT-HANDED
•SUGAR-PHOSPHATE
- Clockwise turn BACKBONE (inside diameter):
11A (1.1nm)
LEFT-HANDED
•BASES (point of
- counterclockwise turn attachment)/A-T and G-C : 11A
(1.1nm)
•Outside diameter of the helix:
20A (2nm)
•COMPLETE TURN of the helix:
34A (3.4nm) contains 10 base
pairs
pH and CHARGES (under
physiological pH)
• Neutral pH: phosphate group
carries a negative charge
(pH7.40)
“Z-DNA”
• formed under conditions of
high salt or in the presence of
alcohol.
• occur in nature when there
is a sequence of alternating
purine pyrimidine.
• longer and narrower than B-
DNA and is a left-handed helix
• the backbone formed a zig-
zag structure.

• Positively charged ions and • repeat helix is 45.6 A˚ and

polypeptides with positively each helical turn has 12 bp
charged side chains must be
associated with DNA in order MINOR GROOVE deep and
to neutralize the negative narrow
charges. MAJOR GROOVE: shallow

“A-DNA”

• observed when DNA is

dehydrated or under high salt
conditions.

• An important shared feature

of A-DNA and B-DNA is that
both are right-handed helices.

• A-DNA is both shorter and

thicker than B-DNA.

• Each repeat double helix in

A-DNA is 24.6 A , and each
turn has about 11 base pairs
(bp).
VARIOUS FORMS OF RNA
Cellular RNA, are almost
always single stranded.
• Nitrogen Bases:
 Adenine
 Guanine
 Cytosine
 Uracil
• 3 MAJOR CLASSES OF RNA
MOLECULES:
1. ribosomal RNA (rRNA)
2. transfer RNA (tRNA)
3. messenger RNA (mRNA)
3 MAJOR CLASSES OF
RNA
rRNA (ribosomal RNA)
-Important structural
components of ribosome,
which functions as non-specific
sites of protein synthesis
during translation.
• Makes up ribosomes
mRNA (messenger RNA)
Carry genetic information from
the DNA of the gene. (protein
encoding)
• The mRNAs vary in size,
reflecting the range in the
sizes of the proteins encoded
by the mRNA.
• Starts in nucleus and moves
to ribosome.
tRNA (transfer RNA)
• It carries amino acids to the
ribosome during translation,
aiding in the translation of
DNA to mRNA to protein.
PHYSICAL PROPERTIES OF
NUCLEIC ACIDS
DENATURATION
• separate two strands of DNA
RENATURATION
• base pair the two strands
together, without disrupting
the covalent bonds that make
up the sugar-phosphate
backbone.
Factors that can affect
denaturation/renaturation:

1. High temperature

2. Low salt concentration

3. High pH (in vitro)

 DNA can be stored short

term at 4°C and long term at
−20°C or colder. RNA is best
stored −20°C or colder.

 Separation of leukocytes is
recommended to avoid
hemoglobin released upon
hemolysis of RBCs.

 Tissue types differ in

stability and nuclease content.

 Nucleic acid from cultured

organisms is best isolated
immediately on harvesting
fresh cultures.

 RNA status depends on the

type of the cell or tissue and
the gene under study.

 Depending on gene
expression, adequate RNA may
be isolated within few hours.
Storage of cell lysates in a
stabilizing buffer is best for
maintaining RNA