Biochemistry/Chemistry 440b

Fall 2014
Solutions to Exam 2
1. Lipid shape refers to the relative nearest neighbor spacing required for optimal
thermodynamic interactions between the non-polar alkyl chains on one hand, and the
polar head groups on the other. When these are matched, the lipid is cylindrical in
shape. This favors the bilayer geometry. If the head group requires a much larger space,
a spherical micelle shape is favored, as is the case for many detergents. If the head groups
require a small space between neighbors, as when they interact with each other, the
hexagonal micelle geometry is preferred.
2. A colligative effect is one that under ideal conditions depends only on the number
concentration of a solute (molarity or molality) and not on the type or molecular size of
the solute. Osmotic pressure, boiling point elevation, freezing point depression and vapor
pressure are all colligative effects. Non-ideal effects often modulate the measurements.
3. The DNA cooperative unit is about 300 or so basepairs. When DNA melts, it does so in
units of about 300 basepairs that form bubbles or loops in the DNA. These units melt in
all-or-nothing fashion, i.e., cooperatively.
4.
a)

b) The unfolded protein has a larger solvent accessible non-polar surface area, which
results in ordered water molecules adjacent to this surface. It takes extra heat to break
these non-covalent bonds between water molecules as the temperature is raised.
Hence, the value of CP is larger for the unfolded protein than for the folded protein,
which does not have as much organized water around non-polar surfaces.
c) The definition of the heat capacity (constant pressure) is

= CP
T P
From this it follows that

= CP
T P
Therefore: H T2 = H T1 + CP (T2 T1 )
This means that the positive value of CP means that the value of H will increase as the
temperature is increased (T2 > T1 )
d) The vant Hoff enthalpy measures the heat per mol of the cooperative unit undergoing
the transition (protein unfolding). Calorimetric enthalpy is normalized per mol of
whatever was placed in the sample compartment.
o
o
Since H cal
, it is plausible that the cooperative unit consists of half of the
2H vH
=

protein. Perhaps the protein has two similar or identical domains that unfold separately
as cooperative units.
5.
a) Start with the following equation:
Tm

o
]
B + [H XY

R ln(CT / 4) + [ A + S

o
XY

+ 11.7 log[ Na + ]
]

We know that the total strand concentration is CT = 50 M and we need to calculate the enthalpy
and entropy terms from the Table of basepair parameters. These are tabulated for log[ Na + ] = 1M
.
For the oligonucleotide duplex:
5- A C G G T G T 3
3- T G C C A C A 5
From the Table of enthalpy and entropy values:
0
0
0
0
0
0
0
0 ]

=
+ [
+
+
+
+
+
0
+
0.2+[-8.8-10.6-8.0-8.4-8.4-8.4] +4.4= -48 kcal/mol
H o =

0
0
0
0
0
0
0
0 ]
0
+

=
+ [
+
+
+
+
+
0
+

o
Soligo
= -5.7+[-22.7-27.2-19.9-22.4-22.4-22.4]-0.0 +13.8= -128.9 cal/(mol-deg)
o
0 because the oligonucleotide is not self-complementary
S sym
=

Now we can find the melting temperature at 1 M NaCl.

0

48000
=
=

0.00005
0
+
128.9
1.98
ln

4
4

=
TM 317.4
=
K 44.3o C

b)
Now we can correct for 0.05M [NaCl]:
+

log[+ ]
+

= 11.7 .

Thus TM2 - TM1=11.7(log[Na ]2 - log[Na ]1)=11.7log([Na ]2/[Na ]1)

at 0.05M [NaCl]:
TM2 = TM1 +11.7log([Na+ ]2/[Na+ ]1)= 44.3+11.7log(0.05)= 44.3 -15.2= 29.2o C
c) By reducing the DNA concentration, the TM will decrease. This can be
seen by looking at the equation, or just by realizing that by mass action,
lowering the concentration of double strand DNA will shift the
equilibrium to favor the single strands:
strand 1 + strand 2 [strand 1strand 2]

6. Relaxed form means no supercoils and all B DNA double strand. We also
know that L=
Tw + Wr .
k
a)
8000
= 762
10.5
Wr = 0

Tw
=

Lk = 762

b) The linking number must remain the same since the plasmid remains
sealed. The twist is reduced by removing 200 basepairs of B DNA and
converting it to single strand. The underwinding of the DNA must be
offset by the formation of positive supercoils.
Lk = 762
8000 200
= 743
10.5
Wr = 762 743 = 19

=
Tw

c) Adding the topoisomerase reduces the writhing number to zero, but the
twist remains the same.
Wr = 0
8000 200
= 743
10.5
Lk = 743

=
Tw

d) Removing the topoisomerase locks the linking number at 743. Removing

the single-strand binding protein results in the entire plasmid being BDNA. This introduces negative supercoils since the linking number is
less than the twist number. There are 19 negative supercoils that result.
Lk = 743
8000
= 762
10.5
Wr =
743 762 =
19

=
Tw

7. Stability is defined as GUwater GUo . We get this from the equation,

recognizing that at 50% unfolded, GUurea =
0 .
GUurea =
GUwater m[urea ]

For protein 1:

0=
GUwater 1.9(3.9)
7.41 kcal/mol
GUwater =
For protein 2:

0=
GUwater 2.1(3.9)
8.19 kcal/mol
GUwater =
o
a) Protein 2 is more stable than protein 1 by 8.19 7.41 =
0.78 kcal/mol at 25 C
b) The two proteins do not necessarily have the same TM values. The TM
depends on the values of HUo and SUo which we do not know.