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Transformation, Transfection,
Chargaff’s Rules and the
Double Helix
History
Bacterial Transformation is
Mediated by DNA
2 Transformation
• T2 Bacteriophage studies
– Bacteriophage = viruses that infect bacteria
– Major chemical components = DNA and protein
– Escherichia coli infected with T2 produce thousands of
new viruses in the host cell
• Host cell lyses and phage are released
Transformation
• The Race is On
– Linus Pauling
– Maurice Wilkins and Rosalind Franklin
– Watson and Crick
• X-ray Crystallography defined
– Diffracted X-rays as they pass through a crystallized
substance
– Patterns of spots are translated by mathematical
equations to define 3-D shape
X-ray Diffractometer
X-Ray Crystallography
• Putting the hydrophobic nitrogenous bases on the inside, and the sugar-
phosphate groups on the outside was a stable arrangement
• Components
•Sugar
•Base
•Phosphate
• 5’ to 3’ direction
• T->U in RNA
• RNA - extra –OH at 2’
of pentose sugar
• DNA deoxyribose
The 5 Bases Purines
Pyrimadines
BASE-PAIRINGS
H-bonds
G C
T A
DNA Double Helix
5 O 3
3 O 5
P P
5 O
1
G C 2 3
4 4
2 1
3 5
O
P P
3
5
O
T A
O 5
P 3 P
The diameter of the helix is 2 nm or 20 Å
2 nm
Geometry of
Watson Crick
Base Pairs
• A:T and G:C pairs are
spatially similar
• 3 H-bonds vs 2
• Sugar groups are attached
asymmetrically on the same
side of the pair
• Leads to a major and
minor grove
• Bases are flat but the
hydrogen bonding leads to
considerable flexibility
Backbone
Conformation
TAUTOMERIC FORMS OF
BASES
• TWO POSSIBILITIES
– KETO (LACTAM)
– ENOL (LACTIM)
• IMPORTANT IN ORDER TO SPECIFY
HYDROGEN BONDING RELATIONSHIPS
• THE KETO FORM PREDOMINATES
TAUTOMERIC FORMS OF BASES
KETO ENOL
Definition of Major
and Minor Groove
Hydrogen
bonding of WC
base pair
Mechanisms of
recognition The canonical Watson-Crick base pair, shown as the G-C pair. Positions of the
minor and major grooves are indicated. The glycosidic sugar-base bond is shown
by the bold line; hydrogen bonding between the two bases is shown in dashed
lines.
MAJOR AND MINOR
GROOVES
• MINOR
– EXPOSES EDGE FROM WHICH C1’ ATOMS
EXTEND
• MAJOR
– EXPOSES OPPOSITE EDGE OF BASE PAIR
• THE PATTERN OF H-BOND POSSIBILITIES IS MORE
SPECIFIC AND MORE DISCRIMINATING IN THE
MAJOR GROOVE
– STUDY QUESTION: LOCATE ALL OF THE
POSSIBILITIES FOR H-BONDING IN THE MAJOR
AND MINOR GROOVES FOR THE 4 POSSIBLE
BASE-PAIRS
STRUCTURE OF
THE DOUBLE HELIX
major
groove
12 Å
one
helical
turn
34 Å minor
groove
6Å
DNA