Nucleic Acid Structure and Hybridisation

Question 1

What are the base pairs structures?

Answer 1

Draw the structures of the base pairs at pH 4, pH 7 and pH 11 (only nucleobases)

Question 2

What is the T_m of a duplex?

Answer 2

The T_m of a duplex is the melting temperature of the duplex.

If the Tm is exceed, the hydrogen bonds between the duplex are broken, and the duplex separates.

Question 3

How do salts affect the melting temperature?

Answer 3

DNA needs cations to shield the charge of the phosphate groups.

Increasing the concentration of the salts increases the melting temperature.

M²⁺ cations are more strongly stabilising than M⁺ cations, and so they increase the melting temperature much more greatly at lower concentrations, although increasing the concentration has little effect on the melting temperature.

Question 4

What is the effect of pH on hybridisation?

Answer 4

• At a neutral pH, the hydrogen bonds between the base pairs are formed normally.
• At pH < 5, the N–H hydrogen bonds denature and the nitrogen atoms are then protonated.
• At pH > 10, the hydrogen bonds are denatured between the Nitrogen and hydrogen, and the nitrogen’s are deprotonated, leaving them with a negative charge.

Question 5

Why does double stranded DNA form a helical structure?

Answer 5

1. Negatively charged phosphate groups repel each other.
2. Hydrogen bonds form between the base pairs.
3. Nucleobases stack on top of eachother via Van der waals forces; this compacts the duplex vertically. This stops water from getting inside.