DNA Structure
Nucleic Acids
DNA and RNA are nucleic acids, long, thread-like polymers
made up of a linear array of monomers called nucleotides
All nucleotides contain three components:
1. A nitrogen heterocyclic base
2. A pentose sugar
3. A phosphate residue
Chemical Structure of DNA vs RNA
Ribonucleotides have a 2’-OH
Deoxyribonucleotides have a 2’-H
Bases are classified as Pyrimidines or Purines
Structure of Nucleotide Bases
The nucleus contains the cell’s DNA (genome)
RNA is synthesized in the nucleus and exported
to the cytoplasm
Nucleus
Cytoplasm
replication
DNA
transcription
RNA (mRNA)
translation
Proteins
Deoxyribonucleotides found in DNA
dA
dG
dT
dC
Nucleotides are
linked by
phosphodiester
bonds
DNA is double stranded
Bases form a specific hydrogen bond pattern
Properties of a DNA double
helix
The strands of DNA are antiparallel
The strands are complimentary
There are Hydrogen bond forces
There are base stacking interactions
There are 10 base pairs per turn
DNA is a Double-Helix
Transcription of a DNA
molecule results in a mRNA
molecule that is singlestranded.
RNase P M1 RNA
RNA molecules do not have a
regular structure like DNA.
hairpin
Structures of RNA molecules
are complex and unique.
RNA molecules can base pair
with complementary DNA or
RNA sequences.
G pairs with C, A pairs with U,
and G pairs with U.
bulge
internal loop
Forces between proteins and DNA
Electrostatic : Salt bridges
Dipolar : Hydrogen bonds
Entropic : The hydrophobic effect
Dispersion : base stacking
Nucleic Acids interact
reversibly with:
Water
Metal ions
Small organic molecules
Drugs
Carcinogens
Antibiotics
Proteins
Major
Groove
Minor
Groove
Groove
Interactions
in DNA
Figure 7.4
Binding to nucleotides in the major groove
The majority of the interactions between proteins and DNA are
hydrogen bonds with functional groups in the major groove
of the double-stranded DNA molecule. Each DNA binding
protein recognizes specific sequences in the DNA.
Hydrogen bonding with N6 and N7 of Adenine, O6 and N7 of Guanine,
O4 of Thymine, and N4 of Cytosine is possible.
Forces between proteins and DNA
Electrostatic : Salt bridges
Dipolar : Hydrogen bonds
Entropic : The hydrophobic effect
Dispersion : base stacking
Forces between proteins and DNA
Electrostatic : Salt bridges
Interaction between groups of opposite charge
Occur between the ionized phosphates of the nucleic acid and either the
e-amino group of lysine, the guanidinium group of arginine, or the
protonated imidazole of histidine.
Forces between proteins and DNA
Dipolar : Hydrogen bonds
d- d+
d- d+
X – H ----- Y – R
X and Y are nitrogen and oxygen in biological systems
Positioning of hydrogen bond donors (X) and acceptors (Y) is
optimized between protein and DNA.
DNA binding proteins contain amino acids that hydrogen
bond to functional groups in the major groove of DNA
Forces between proteins and DNA
Entropic : The hydrophobic effect
A complementary surface formed between a protein and a
nucleic acid will release ordered water molecules at the surface of
the protein or nucleic acid.
The formerly ordered water molecules become part of the
disordered bulk water, thus stabilizing the interaction through
an increase in the entropy of the system.
Consequently, the surfaces of the protein and nucleic acid tend to
be exactly complementary, increasing the specificity of the interaction.
Forces between proteins and DNA
Dispersion : base stacking
Base stacking is dependent on the hydrophobic effect as
well as dispersion (London) forces.
Molecules with no net dipole can attract each other by a transient
dipole-induced dipole effect.
These forces are weak but do play a role in
protein – nucleic acid interaction,
specifically in base stacking.
E. coli DNA polymerase III has a doughnut-shaped hole lined
with positively-charged amino acid side chains that
interact with the negatively-charged DNA strand
a-helix
The catabolite activator protein
(CAP) from E. coli uses alpha
helices to interact with
nucleotide bases in the major
groove of the DNA helix.
An amine-containing amino acid side
chain often forms hydrogen bonds with
major groove bases.
A single amino acid may form hydrogen
bonds with multiple, adjacent
nucleotide bases, increasing sequencespecific interaction.
Common amino acid: arginine or
glutamine
a-helix
The kinetics of forming protein – DNA complexes
Proteins often bind to specific sequences of DNA.
Example: Restriction enzyme EcoRI binds to the DNA sequence
5’-GAATTC-3’
3’-CTTAAG-5’
How do proteins find their target DNA sequence?
1. Randomly bind, dissociate, re-bind until they find their sequence?
(Three-dimensional random walk)
2. Bind non-specifically and then slide along DNA until they find it?
(One-dimensional walk)
Non-sequence specific protein – DNA
interaction
From the moment a new strand of DNA is synthesized to the moment
it is degraded in a cell, there are proteins associated with it.
Many of these proteins interact in a non-sequence specific manner.
Many of the proteins are involved in packaging the DNA.
Example: histone proteins that form the nucleosome
Proteins that interact non-specifically with DNA interact with the
negatively-charged ribose-phosphate backbone. Therefore, they have
a high percentage of basic amino acid side chains such as lysine and
arginine.
DNA in eukaryotic cells is packaged into nucleosomes,
which contain proteins called histones.
Nucleosomes
DNA wrapped around a histone core (side view)
Specific protein – DNA interactions
For a cell to function, proteins must distinguish one
nucleic acid sequence from another very accurately.
Activators and repressors of transcription
turn specific genes on and off.
Common themes of protein - DNA interaction
1. Helix-turn-helix
2. Zinc finger
3. Leucine zipper
Helix-turn-helix motif
Often found in proteins that regulate gene transcription.
The pair of a-helices stack to form a V shape with an angle of about 60º
The first helix positions the second helix.
The second helix binds to the DNA, projecting into the major groove
and recognizing specific sequences.
Shown is a
helix-turn-helix motif
from a homeodomain protein,
A family of proteins that
binds to eukaryotic DNA
and regulate transcription
of specific genes.
Zinc finger motif
Often found in proteins that regulate gene transcription.
A zinc is coordinated to cysteine or histidine residues of the protein.
An a-helix is inserted into the major groove and binds DNA.
histidine
histidine
Shown is a
zinc finger motif
from a repressor protein
from a phage, a
bacterial virus
zinc
cysteine
cysteine
Zinc finger motif
Often found in proteins that regulate gene transcription.
A zinc is coordinated to cysteine or histidine residues of the protein.
An a-helix is inserted into the major groove and binds DNA.
cysteine
cysteine
Shown is a
zinc finger motif
from the glucocorticoid
receptor, a protein that
mediates hormone action.
Two zincs are present.
zinc
Leucine zipper motif
Often found in proteins that regulate gene transcription.
Two alpha helices interact through interaction between hydrophobic
leucine amino acid side chains on one side of the alpha helix.
Shown is a
leucine zipper
protein
Negative regulatory proteins bind to operator sequences
in the DNA and prevent or weaken RNA polymerase binding
Most prokaryotic mRNA molecules are polycistronic, they
encode multiple genes. These genes are usually
involved in the same biochemical event.
A single promoter controls the expression of these genes.
This functional unit of DNA is called an operon.
A classical example of
transcriptional regulation
is lactose metabolism
in E. coli.
Proteins required for
lactose metabolism in
E. coli are encoded by
the lac operon.
The E. coli lac operon
lacI – encodes the Lac repressor protein
lacZ – encodes b-galactosidase
lacY – encodes galactose permease
lacA – encodes transacetylase
O2 and O3 are pseudooperators
The Lac repressor protein is thought to bind to the main operator
and one of the pseudooperators, forming a loop in the DNA.
When lactose is present in high concentrations, the lactose metabolism
gene products are needed in a cell. In the absence of lactose, the Lac
repressor protein binds to the operator in the DNA, repressing
transcription. The Lac repressor, however, binds to allolactose, a
metabolite of lactose, inducing a conformational change that abolishes
binding to the DNA operator sequence.
Transcription is no longer repressed.
- allolactose  transparent
+ allolactose  bold
DNA binding proteins contain amino acids that hydrogen bond to
functional groups in the major groove of DNA.
DNA sequences recognized by regulatory proteins are often inverted repeats of a short
DNA sequence. These repeats form a palindrome with two-fold symmetry about a central
axis. Regulatory proteins are often dimeric. Each subunit binds to one strand of the DNA.
5’-TACGGTACTGTGCTCGAGCACTGCTGTACT-3’
3’-ATGCCATGACACGAGCTCGTGACGACATGA-5’
central axis
The Lac repressor protein
The Lac repressor is a
tetramer of four identical
protein subunits.
There are DNA-binding
domains on each subunit
shown in blue.
The allolactose binding
domain (green) is connected
to the DNA binding domain
through linker helices (yellow).
Tetramerization domains (red)
form contacts between subunits.
The Lac repressor protein
The Lac repressor is a
tetramer of four identical
protein subunits.
There are DNA-binding
domains on each subunit
shown in blue.
The allolactose binding
domain (green) is connected
to the DNA binding domain
through linker helices (yellow).
Tetramerization domains (red)
form contacts between subunits.
The DNA binding domains of the Lac repressor contain a
helix-turn-helix motif, a structure critical for the interaction of many
proteins with DNA.
helix
turn
helix
Lac repressor
protein
(lacI)
Figure 8-21
Lac repressor bound to DNA
Figure 8-22
Lac repressor bound to DNA
Figure 8-23
Protein – DNA interactions
Figures 8-16 and 8-17
Transcriptional elements of a
eukaryotic structural gene
Figure 9.1 page 151
Transcriptional Activation
Figure 9.2 page 152
TATA box sequences
Structure of
the TATA box
binding
protein
Figure 9.4 page 155
Structure of TBP complexed with DNA
Figure 9.5 page 156
DNA bound to TBP is bent
Figure 9.6 page 156
Sequence
specific
interactions
between TBP
and DNA
Figure 9.7 page 157
Transcription Factors
Chapter 10
Helical wheels of DNA-binding
domains of transcription factors
Figure 10.17 page 192
Regulatory proteins that function as dimers contain
regions of amino acid sequence that mediate interaction
between protein subunits.
One common motif is the leucine zipper.
5’-TACGGTACTGTGCTCGAGCACTGCTGTACT-3’
3’-ATGCCATGACACGAGCTCGTGACGACATGA-5’
central axis
Fig 28-14
The leucine residues of a leucine zipper provide
hydrophobic interaction between alpha helices at regular intervals.
Fig 28-14
Leucine Zipper
Figure 10.18 page 193
Heterodimerization of
leucine zipper proteins
Figure 10.19 page 193
Transcription Factor GCN4
Figure 10.20 page 194
DNA-binding domain of GCN4
Figure 10.21 page 195
GCN4-DNA interactions
Figure 10.22 page 195