Another important feature of RNA (or DNA, for that matter) is its base
composition, that is, the percentage of each of the nucleotides that
make it up. The four bases that, with ribose and phosphoric acid,
comprise the RNA molecule are guanine, adenine, cytosine, and uracil. It
will be noted that three of the four—guanine, adenine, and cytosine—are
the same as those in DNA, but thymidine in DNA has been replaced by
another base, uracil. To determine the percentage of each base in a
given RNA molecule, we must digest RNA with alkali to produce
mononucleotides, which are smaller molecules, each consisting of a base,
ribose, and phosphoric acid. We can now separate the four nucleotides by
using paper chromatography (see Figure 20).
[Illustration: Figure 21 _A paper chromatography showing separation of
amino acids in two directions. Radioactivity in samples then produced
this record by radioautography._]
In this technique a mixture of compounds is deposited on the edge of a
special type of paper. This edge is then immersed in a solvent that
slowly permeates the paper (at a constant speed) by capillary action. As
the solvent moves from the immersed edge toward the other edge, which is
hanging freely, it carries the mixture of nucleotides with it. Each of
the compounds in the mixture travels at a different speed, however;
thus, as the solvent front moves along the paper, the dissolved
compounds are separated from each other and appear as distinct spots on
the paper. To locate the nucleotides on the paper and to determine the
percentage composition, we can use a chromatogram scanner, a device that
scans the paper chromatograms, measures the radiation from them, and
thus locates the labeled substances (see Figure 22).
[Illustration: Figure 22 _Recording of radioactivity in a sample by
radioautography and paper chromatography. The peaks of the trace
prepared by a chromatogram scanner coincide with the areas of separated
components on the same chromatogram, as revealed by radioautography. The
radioautograph is superimposed on the chromatogram recording._]
Another technique used to separate the nucleotides of RNA is column
chromatography. In this method mixtures of nucleotides are separated as
they pass down a column of chemicals (see Figure 23).
[Illustration: Figure 23 _Students visiting Argonne National Laboratory
listen to a scientist explain the column chromatography process, in
which mixtures of nucleotides are separated as they pass down a column
of chemicals._]
We have now learned how to use radioisotopes to investigate the
synthesis of RNA, the molecule that translates the DNA message into the
language of proteins. Let us now see what we can learn about the
synthesis and function of proteins.