DNA Structure. All nucleotides contain three components: 1. A nitrogen heterocyclic base 2. A...
Preview:
Citation preview
- Slide 1
- DNA Structure
- Slide 2
- 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
- Slide 60
- Transcription Factor GCN4 Figure 10.20 page 194
- Slide 61
- DNA-binding domain of GCN4 Figure 10.21 page 195
- Slide 62
- GCN4-DNA interactions Figure 10.22 page 195