Question

Which conformation of DNA, totally double helix, minimally unwound or largely unwound, would have the highest relative optical density at \(260 \mathrm{nm}\) ? Would a molecule of DNA having a higher content of guanine and cytosine than of adenine and thymine have a higher or lower \(T_{m}\) than one with the reverse composition?

Short answer

Answer: The largely unwound DNA conformation has the highest relative optical density at 260 nm. A DNA molecule with a higher content of guanine and cytosine would have a higher melting temperature than one with more adenine and thymine.

Step-by-step solution

Understanding DNA Conformation

DNA exists in different conformations, depending on the environmental conditions and molecular interactions. The three conformations mentioned in the exercise are the totally double helix, minimal unwound, and largely unwound, which refer to the degree of separation of the two DNA strands. A fully double helix structure is when the DNA strands are tightly wound together and form the classical double helix structure. Minimal unwound occurs when minimal base pair separation happens, and the structure retains its overall double helix shape. Largely unwound occurs when a significant amount of base pair separation happens, but the strands still remain together.

Relative Optical Density

Optical density, or absorbance, is a measure of how much light is absorbed by a substance as it passes through it. In the case of DNA, absorbance or relative optical density at 260 nm can be used to estimate the concentration of DNA in a solution because the nitrogenous bases absorb ultraviolet (UV) light.

Identifying the Highest Optical Density Conformation

Among the three conformations, the largely unwound DNA would have the highest relative optical density at 260 nm. This is because when the DNA is more unwound, more nitrogenous bases are exposed and available to absorb the 260 nm light. In the totally double-helix structure, the bases are more internally stacked and less accessible to the light, causing lower absorbance.

Melting Temperature (\(T_m\))

The melting temperature (\(T_m\)) of DNA is the temperature at which half of the DNA molecules are denatured—meaning, the double helix becomes separated into single strands. The \(T_m\) is influenced by the base pair composition of the DNA molecule, with the GC content being a crucial factor. Guanine (G) and cytosine (C) have three hydrogen bonds between them, while adenine (A) and thymine (T) have only two.

Comparing \(T_m\) of Different Base Pair Compositions

A DNA molecule with a higher content of guanine and cytosine would have a higher \(T_m\) than one with more adenine and thymine. This is due to the extra hydrogen bond between guanine and cytosine making it more stable and requiring more energy, or a higher temperature, to break the base pairs apart and denature the DNA molecule. In conclusion, the largely unwound DNA would have the highest relative optical density at 260 nm, and a DNA molecule with a higher content of guanine and cytosine would have a higher melting temperature than one with more adenine and thymine.