DNA Structure. All nucleotides contain three components: 1. A nitrogen heterocyclic base 2. A...
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DNA Structure
DNA Structure. All nucleotides contain three components: 1. A nitrogen heterocyclic base 2. A pentose sugar 3. A phosphate residue Nucleic Acids DNA and
All nucleotides contain three components: 1. A nitrogen
heterocyclic base 2. A pentose sugar 3. A phosphate residue Nucleic
Acids DNA and RNA are nucleic acids, long, thread-like polymers
made up of a linear array of monomers called nucleotides
Slide 3
Ribonucleotides have a 2-OH Deoxyribonucleotides have a 2-H
Chemical Structure of DNA vs RNA
Slide 4
Structure of Nucleotide Bases Bases are classified as
Pyrimidines or Purines
Slide 5
The nucleus contains the cells DNA (genome) RNA is synthesized
in the nucleus and exported to the cytoplasm Nucleus Cytoplasm DNA
RNA (mRNA) Proteins replication transcription translation
Slide 6
dA dG dTdC Deoxyribonucleotides found in DNA
Slide 7
Nucleotides are linked by phosphodiester bonds
Slide 8
Bases form a specific hydrogen bond pattern DNA is double
stranded
Slide 9
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 Properties
of a DNA double helix
Slide 10
DNA is a Double-Helix
Slide 11
RNase P M1 RNA Transcription of a DNA molecule results in a
mRNA molecule that is single- stranded. RNA molecules do not have a
regular structure like DNA. 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 hairpin
Slide 12
Electrostatic : Salt bridges Dipolar : Hydrogen bonds Entropic
: The hydrophobic effect Dispersion : base stacking Forces between
proteins and DNA
Slide 13
Water Metal ions Small organic molecules Drugs Carcinogens
Antibiotics Proteins Major Groove Minor Groove Nucleic Acids
interact reversibly with:
Slide 14
Groove Interactions in DNA Figure 7.4
Slide 15
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 N 6 and N 7 of
Adenine, O 6 and N 7 of Guanine, O 4 of Thymine, and N 4 of
Cytosine is possible. Binding to nucleotides in the major
groove
Slide 16
Electrostatic : Salt bridges Dipolar : Hydrogen bonds Entropic
: The hydrophobic effect Dispersion : base stacking Forces between
proteins and DNA
Slide 17
Electrostatic : Salt bridges Interaction between groups of
opposite charge Occur between the ionized phosphates of the nucleic
acid and either the -amino group of lysine, the guanidinium group
of arginine, or the protonated imidazole of histidine. Forces
between proteins and DNA
Slide 18
Dipolar : Hydrogen bonds - + - + 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. Forces between proteins and DNA
Slide 19
DNA binding proteins contain amino acids that hydrogen bond to
functional groups in the major groove of DNA
Slide 20
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
Slide 21
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. Forces between proteins and DNA
Slide 22
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
Slide 23
-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 sequence- specific interaction. Common amino acid:
arginine or glutamine
Slide 24
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) The
kinetics of forming protein DNA complexes
Slide 25
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. Non-sequence specific protein DNA interaction
Slide 26
Nucleosomes DNA in eukaryotic cells is packaged into
nucleosomes, which contain proteins called histones. DNA wrapped
around a histone core (side view)
Slide 27
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 Specific protein DNA interactions
Slide 28
Often found in proteins that regulate gene transcription. The
pair of -helices stack to form a V shape with an angle of about 60
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. Helix-turn-helix
motif
Slide 29
Often found in proteins that regulate gene transcription. A
zinc is coordinated to cysteine or histidine residues of the
protein. An -helix is inserted into the major groove and binds DNA.
Shown is a zinc finger motif from a repressor protein from a phage,
a bacterial virus zinc histidine cysteine Zinc finger motif
Slide 30
Often found in proteins that regulate gene transcription. A
zinc is coordinated to cysteine or histidine residues of the
protein. An -helix is inserted into the major groove and binds DNA.
Shown is a zinc finger motif from the glucocorticoid receptor, a
protein that mediates hormone action. Two zincs are present. zinc
cysteine Zinc finger motif
Slide 31
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 Leucine zipper motif
Slide 32
Negative regulatory proteins bind to operator sequences in the
DNA and prevent or weaken RNA polymerase binding
Slide 33
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.
Slide 34
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.
Slide 35
The E. coli lac operon lacI encodes the Lac repressor protein
lacZ encodes -galactosidase lacY encodes galactose permease lacA
encodes transacetylase O 2 and O 3 are pseudooperators
Slide 36
The Lac repressor protein is thought to bind to the main
operator and one of the pseudooperators, forming a loop in the
DNA.
Slide 37
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
Slide 38
DNA binding proteins contain amino acids that hydrogen bond to
functional groups in the major groove of DNA.
Slide 39
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
Slide 40
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.
Slide 41
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.
Slide 42
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
Slide 43
Lac repressor protein (lacI) Figure 8-21
Slide 44
Lac repressor bound to DNA Figure 8-22
Slide 45
Lac repressor bound to DNA Figure 8-23
Slide 46
Protein DNA interactions Figures 8-16 and 8-17
Slide 47
Transcriptional elements of a eukaryotic structural gene Figure
9.1 page 151
Slide 48
Transcriptional Activation Figure 9.2 page 152
Slide 49
TATA box sequences
Slide 50
Structure of the TATA box binding protein Figure 9.4 page
155
Slide 51
Structure of TBP complexed with DNA Figure 9.5 page 156
Slide 52
DNA bound to TBP is bent Figure 9.6 page 156
Slide 53
Sequence specific interactions between TBP and DNA Figure 9.7
page 157
Slide 54
Transcription Factors Chapter 10
Slide 55
Helical wheels of DNA-binding domains of transcription factors
Figure 10.17 page 192
Slide 56
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. Fig 28-14
5-TACGGTACTGTGCTCGAGCACTGCTGTACT-3
3-ATGCCATGACACGAGCTCGTGACGACATGA-5 central axis
Slide 57
The leucine residues of a leucine zipper provide hydrophobic
interaction between alpha helices at regular intervals. Fig
28-14
Slide 58
Leucine Zipper Figure 10.18 page 193
Slide 59
Heterodimerization of leucine zipper proteins Figure 10.19 page
193
