Download Five types of bases.

Structure and function of nucleic
acids.
Heat.
Heat flows through the boundary of the system because there exists a
temperature difference between the system and surroundings.
dq  Cdt
C – heat capacity, can be measured by putting known amount of heat
into the system and measuring the T-difference.
Work.
Pex
x
gas
P-V work:
dw   Pex Adx   Pex dV
P
P1,V1
P3,V3
P2,V2
V
The first law of thermodynamics.
The energy of isolated system is constant:
E  q  w
E – state function, energy change is the same regardless how this
change was produced:
E2
E   dE  E2  E1
E1
Enthalpy.
At constant pressure:
H  E  PV
Calculation of total enthalpy of reactions. The enthalpy is a state function, it
can be added and subtracted for a sequence of reactions:
AB (ΔH1); AC (ΔH2); CD (ΔH3);DB (ΔH4).
ΔH1= ΔH2+ ΔH3+ ΔH4
Entropy.
What defines the direction of spontaneous change?
Statistical aspect of entropy:
Ex: rearranging three balls:
123 132 213 231 312 321
Spontaneous process will go in the direction of increasing the probability W.
S  K ln W
S  nR ln( V f / Vi )
The second law of thermodynamics.
It shows how much heat should be used to make the system
disordered given an existing level of disorder measured by T.
dS  dqrev / T
For irreversible change in universe:
S  S gas  S sur  nR ln( V f / Vi )  0
Free energy.
Gibbs free energy at P= const:
G = H –TS
At equilibrium ΔG = 0 and:
G 0   RT ln K
K – equilibrium constant
Standard state: at T = 25C and P = 1 atmosphere.
DNA structure.
History:
• 1868 Miescher – discovered nuclein
• 1944 Avery – experimental evidence that DNA is
constituent of genes.
• 1953 Watson&Crick – double helical nature of DNA.
• 1980 X-ray structure of more than a full turn of B-DNA.
DNA structure.
• Genomic DNAs are large molecules:
Eschericia coli: 4.7 x 10^6 bp; ~ 1 mm contour
length; Human: 3.2 x 10^9 bp; ~ 1 m contour
length
• Some DNA molecules (plasmids) are circular
and have no free ends (mtDNA, bacterial DNA)
• Average gene of 1000 bp can code for average
protein of about 330 amino acids
Five types of bases.
Nucleotides and phosphodiester bond.
Phosphodiester bond
Complementarity of nucleosides – bases
for double stranded helical structure.
Chargaff’s rule.
• Amounts of G = C, A = T.
• Most DNAs obey this rule – double starnded,
few RNAs obey this rule – single stranded.
• Why RNAs do not reach the maximum of base
pairing?
- RNA contains not equal amount of A/U and G/C.
- RNA contain modified bases which prevent base
pairing
Double helical structure of DNA.
A- and B-DNA – right-handed helix,
Z-DNA – left-handed helix
B-DNA – fully hydrated DNA in vivo,
10 base pairs per turn of helix
Major and minor grooves of DNA.
• The sugar-phosphate backbones
are bulky and form ridges on the
edges of the helix, between
these edges the part of bases
are exposed forming grooves.
• Base pairs are tilted and this
arrangement makes the major
groove very deep and minor
groove very shallow.
Hydration of B-DNA.
From R. Dickerson, Structure & Expression
Major and minor grooves of DNA.
Copyright © Ramaswamy H. Sarma 1996
Sequence specific recognition of DNA by
proteins.
• Nitrogen and oxygen exposed in the grooves can make
hydrogen bonds with proteins.
• Different Watson/Crick base pairs have different patterns
of donors and acceptors
- H-bond acceptor
- hydrogen atom
- H-bond donor
- methyl group
G
C
G
C
A
T
A
T
C
G
C
G
T
A
T
A
Major groove
Minor groove
DNA thermodynamics.
Two major types of interactions:
• Base pairing (hydrogen bonds)
• Base stacking of nearest neighbors (π-electron sharing
of aromatic rings+ hydrophobic)
G  G
init
 G
pairing
 G
stacking
Thermodynamic parameters for base
pairing and stacking.
Base pairing (kcal/mol):
A–A U–U A–U C–C
-0.67 -1.07 -2.72 -1.97
G–G
-5.00
G–C
-6.00
Base stacking:
5’-A-A -1.4
3’-T-T
5’-A-C
3’-T-G
-1.8
5’-G-A
3’-C-T
-1.7
5’-A-T
3’-T-A
-0.9
5’-C-A
3’-G-T
-1.6
5’-C-G
3’-G-C
-2.5
5’-T-A
3’-A-T
-0.8
5’-A-G
3’-T-C
-1.3
5’-G-C
3’-C-G
-2.5
5’-C-C -2.1
3’-G-G
Helix initiation (at least one GC-pair)
Helix initiation (no GC-pairs)
+1.8
+2.7
Classwork 1.
Calculate ΔG for DNA stabilization:
5’-T-A-C-T-G-3’
3’-A-T-G-A-C-5’
Difference between DNA & RNA:
Differences between DNA & RNA:
• T is replaced by U
• Extra –OH group at 2’ pentose sugar, sugar is ribose, not
deoxyribose
• RNA usually does not form double helix, makes loops
within one strand, often contains modified bases
• RNA has an additional 2’-OH group which can form HB,
stabilizing tertiary structure
RNA functions.
• Information decoding (mRNA)
• Information transfer (tRNA)
• Structural molecule (rRNA)
• Catalytic function (ribozymes)
• Regulatory function
Types of RNA secondary structures.
• hairpin loop, (i ,j) defines a hairpin loop if i and j are
paired and k is a free base, i < k < j.
• stacked loop – if i and j are paired and (i+1) and (j-1) are
paired.
• internal loop – two closing base pairs and all bases
between them are free.
• multi-branched loop or multi-loop – loop has at least
three closing base pairs.
• pseudoknot – (i,j,i’,j’) defines pseudoknot if i and j are
pairs, i’ and j’ are pairs and i < i’ <j < j’.
Illustration of RNA secondary structures.
From M.S. Andronescu
Tertiary structure of rRNA.
Information decoding (mRNA).
Information transfer (tRNA).
Characteristic properties:
- Each tRNA is specific for one of the amino
acids.
- Forms a lot of tertiary interactions (Ex: tRNA
for Phe contains 20 base pairs which form 52
HBs and 40 HBs from tertiary interactions).
- Most of tRNA structure is common between
the species.
Structural molecule (rRNA).
Ribosome of prokaryotes: 3 RNA molecules
and 55 proteins. Small subunit controls tRNA
interactions with mRNA;
large subunit controls catalysis.
RNA catalytic properties: ribozymes.
• RNA of self-splicing group I introns, contain 4
sequence elements and form specific secondary
structures
• RNA self-splicing group II introns
• RNA from viral and plant satellite RNAs
• Ribosomal RNAs
Classwork 2.
1. Go to http://ndbserver.rutgers.edu/.
2. Select Crystal structure of B-DNA, resolution
>=2 Angstroms.
3. Select Crystal structure of single-stranded RNA
with mismatch base pairing with resolution >= 2
Angstroms.
Classwork 3: DNA topoisomerase.
1. Go to http://ndbserver.rutgers.edu/.
2. Select a structure for DNA – topoisomerase complex
3. Look at the complex using CN3D and answer the
questions:
- What type of groove does the topoisomerase bind
to?
- What types of secondary structure are involved in the
interaction between DNA and protein?
RNA secondary structure prediction
Assumptions used in predictions:
- The most likely structure is the most stable one.
- The energy of each base pair depends only on the
energy of the previous base pair.
- Energy parameters for different types of RNA secondary
structures are derived from the experiment.
- The structure is formed w/o knots.
Minimum energy method of RNA
secondary structure prediction.
• Self-complementary regions can be found in a dot matrix
• The energy of each base pair depends only on the
energy of the previous base pair
• Energy parameters for different types of RNA secondary
structures are derived from the experiment
• The most energetically favorable conformations are
predicted by the method similar to dynamic programming
Minimum energy method of RNA
secondary structure prediction.
Classwork II: Predict secondary structure
for RNA “ACGUGCGU”.
Stacking energies for base pairs
A/U
C/G
G/C
U/A
G/U
U/G
A/U
-0.9
-1.8
-2.3
-1.1
-1.1
-0.8
C/G
-1.7
-2.9
-3.4
-2.3
-2.1
-1.4
G/C
-2.1
-2.0
-2.9
-1.8
-1.9
-1.2
U/A
-0.9
-1.7
-2.1
-0.9
-1.0
-0.5
G/U
-0.5
-1.2
-1.4
-0.8
-0.4
-0.2
U/G
-1.0
-1.9
-2.1
-1.1
-1.5
-0.4
Destabilizing energies for loops
Number of
bases
1
5
10
20
30
Internal
-
5.3
6.6
7.0
7.4
Bulge
3.9
4.8
5.5
6.3
6.7
Hairpin
-
4.4
5.3
6.1
6.5
Sequence covariation method.
Some positions from different species can covary because they are
involved in pairing
fm(B1) - frequences in column m;
fn(B2) – frequences in column n;
fm,n(B1,B2) – joint frequences of two nucleotides in two columns.
f m,n ( B1 , B2 ) /( f m ( B1 )  f n ( B2 ))
Seq 1
Seq 2
Seq 3
Seq 4
---G------C-----G------C-----A------T-----T------A---
Ribozymes.
• RNA of self-splicing group I introns, contain 4
sequence elements and form specific secondary
structures
• RNA self-splicing group II introns
• RNA from viral and plant satellite RNAs
• Ribosomal RNAs
GenBank – an annotated collection of all
publicly available DNA sequences.
Human Genome project.