DNA: Structure and Function

Deoxyribonucleic acid (DNA) is how the cell encodes information to make all of the macromolecules that it requires. Most of these macromolecules are proteins but some are RNA molecules. From a chemical point of view DNA is a polymer made up of a ribose and phosphate backbone with varible side groups of Adenine(A), Cytosine(C), Guanine(G), and Thymine(T). Note that the polymer has a directionality associated with it because on one side of the riobse backbone the phosphate is bound to the 5' position and on the other the phosphate is bound to the 3' position, as illustrated in Figure 1. By convention DNA is represented by a string of A, T, G and C written starting at the 5' end and continuing left to right to the 3'end of the molecule. For example the strand on the left hand side of Figure 1 would be written: CAGT. The strand on the right hand side of Figure 1 would be written ACTG.

From R.E. Dickerson, "The DNA helix and how It is Read." Copyright 1983 by Scientific American, Inc.

Figure One: DNA and its complementary strand. The helix has been flattened to facilitate the viewing of the chemical interactions. The ribose phosphate backbone is colored in blue and each side group is a different color. Note that the ribose makes the molecule dirctional as the phosphate in one direction is bound at the 3 prime position and in the other direction the phosphate is bound to the 5 prime position. By convention DNA is written five prime to three prime.

DNA's most amazing characteristic is that it will bind to itself in a very specific manner, forming a helix. Figure 1 illustrates two DNA molecules binding. The variable groups of the polymer interact with eachother to mediate this binding. Adenine interacts with Thymine and Cytosine interacts with Guanine. No other bonds are energetically favorable, for example Cytosine cannot bond with itself. The bonds that these variable groups form are called are called base pairs and are illustrated in Figure 2. Thus, by knowing the sequence of bases of one strand of DNA we immediately know the sequence of the DNA strand which will bind to it, this strand is called the reverse complement or just the complementary strand. In Figure 1 the right hand strand is the reverse complement of the left hand side and vice-versa.

From G. S. Sent, Molecular Biology of Bacterial Viruses. Copyright 1963 by W. h. Freeman and Company

Figure Two:Illustration of DNA basepairing. Thermodynamically the interactions Adenine, Guanine, Cytosine, and Thymine are driven by both the hydrophobic stacking interactions hydrogen bond interactions. The specificity of the base pairing is driven mainly by hydrogen bonding and stearic interference.

This relationship to the complementary strand has many implications that make life as we know it possible, and allows all sorts of molecular biology techniques. In one of the greatest understatements of all time Watson and Crick said it best in their original paper on the structure of DNA:

"It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.".

In the cell DNA is stored in helical form with its complementary strand. When the cell need to duplicate the DNA the strands for cell division the helix is split apart and each strand is used as a template to make a complementary strand of itself, an amazingly elegant solution.

Molecular biology capitalizes in many ways on this process of helix formation. By taking a DNA sequence which is known it is possible to examine large pools of DNA to see if your DNA of iterest is present and how much is present. This is known as a hybridization experiment. This technique makes use of the fact that a DNA sequence will bind specifically to its reverse complement, even in the presence of other molecules. Generically the protocol is as follows:

1. Isolate a pool of DNA sequences of interest, e.g. from a particular strain of bacteria.
2. Run the DNA sequences on a gel to separate them by size.
3. Transfer the DNA sequences from the gel to a membrane, typically nylon.
4. Label the DNA sequence that you have and know the sequence of, usually with radioactive phosphate (Don't worry, it's plenty safe), this is now considered your probe.
5. Place the labeled DNA probe in solution with the membrane which contains your pool of DNA sequences.
6. If your sequence of interest is in the pool of DNA sequences the labeled DNA probe will bind to it and form a stable helix with it.
7. Wash away the non-specific unstable helixes.
8. Look for radioactivity on the blot, the presence of your DNA probe indicates the presence of your sequence of interest in the pool.

It doesn't really matter how you label the DNA, what surface you mount it on, or how you separate it. The basic idea is to use a DNA molecule of known sequence to probe for it's complementary strand in a pool of unknown DNA sequences.

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