Excelet

DNA and Thermal Denaturation

Here are the chemical structures of the bases found in DNA.
NH2

NH2

N
NH

NH

A d e n in e

C y t o s in e

O
N
O

NH

O
NH

G u a n in e

NH2

H3C

NH
NH

T h y m in e

The bases interact in pairs to hold the two helical strands together. Let's examine the interaction
and what happens if we heat a solution of dissolved DNA.
For a good interactive Chime structure of DNA click here
Click on the tabs to navigate!!

Building a short double-strand of DNA

How do the bases pair in the DNA molecule?
Build one strand of DNA with 10 pairs of bases - select either a, c, g, or t.
The Ediss is the energy of dissociation to separate the strands.
place your base choices here
Ediss for
c
g
t
a
g
c
a
t
g
c

your strand

1000
2

g
c
a
t
c
g
t
a
c
g

0
600
5 + 600 750
f(x) = 30x
10
900

2
800
3
3
600
10
Ediss

400
4

240

200
6

540
1

10

number of G-C pairs

complimentary strand
will appear here

30

780

0
0

Hydrogen bond energy

What holds
the strands
together?
answer

Number of A-T pairs

4

Number G-C pairs

6

Energy of dissociation
780
kJ/mole

kJ/mole
What happens to the energy of dissociation as more "g" or "c" is added to the strand? Why?

Sinex 2007
We are assuming that ALL hydrogen bonds between the base pairs have the SAME dissociation energy!
The protection is turned on to avoid messing up formulas in cells.

your strand
What holds
the strands

Calculating the temperature to separate the double strand to single strands - thermal denaturation of DNA
Build one strand of DNA with up to 20 pairs of bases - select either a, c, g, or t.
place your base choices here
How does the length of the nucleotide and
its base composition influence the
temperature of thermal denaturation, Tm?
a
t
t
g
g
c
t
a
g
c
a
t
g

a
c
c
g
a
t
c
g
t
a
c

length of nucleotide =

12

number of G-C pairs =

number of A-T pairs =

%G-C pairs =

50

melting point, Tm =

36

blk
a
t
g
c

8
3
3
4
2

Explore the Tm:

Compare a nucleotide of 6 A-T with 6 G-C.
How about 3 A-T and 3 G-C to the above?
Compare a nucleotide of 6, 12, and 18 A-T.
This is actually the temperature where half the
double strands have separated. It is measured
by a change in UV absorption at 260 nm while
increasing the temperature. A number of
theoretical calculations are available to
predict this temperature.

It is incorrectly referred to as a melting point in biochemistry.

This temperature is the temperature of thermal denaturation.
Sinex 2007

Rank the order of the melting points of the DNA strands

AT
CG
GC
AT
CG
TA
GC
AT
TA
CG
MP rank:

CG
GC
AT
CG
GC
TA
CG

CG
AT
TA
AT
AT
GC
AT
TA
GC
AT

AT
TA
GC
TA
AT
TA
AT
TA

Do you understand the temperature of thermal denaturation, T m?

Which strand has the highest? Which the lowest? Why?

Hint

1 = hightest mp...

Sinex 2010

F9
1

2 3 4 5

6 7 8 9 10 11 12 13 14 15 16 17 18 19###21###
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1 1 3 4 1 3 4 2 4 2 1

1 4 2 4 4 1 4 2 3 3 2 3 3 3 4 2 2 3 1 4 1 4 4 1 3 4 3 2 3 3 4 2 4 1 4 2 2 2 1

A A G C A G C T C T A A C T C C A C T G G T G G G C T T G A C A C C A G C G T G G C T C A C T T T A