Changes in the DNA Base Sequence

Our genetic information is contained in the precise ordering of the DNA base pairs. Although the DNA-copying process is very accurate, rare errors do occur. As in computers that are equipped with an error-checking system, cells possess mechanisms that minimize mistakes in DNA replication. Yet each time our cells divide, all 3 billion base

Figure 7.3 How Karyotypes Are Made. First cells are obtained from blood or other cells of an adult or a fetus through chorionic villus sampling or amniocentesis. If necessary, for example, with adult cells, phytohemagglutinin, a chemical that stimulates growth and division is added. Then, after growth, colcemid, a chemical that stops the cells in mitosis is added. Then these cells are swollen in a low-salt solution and dropped onto a microscope slide. This step bursts open the cells and spreads out the chromosomes. The chromosomes are then stained and photographed through the microscope. In the past, photographs of chromosomes were literally cut up with scissors and arranged by size and shape. Now, this is all done with image-analysis software.

Figure 7.3 How Karyotypes Are Made. First cells are obtained from blood or other cells of an adult or a fetus through chorionic villus sampling or amniocentesis. If necessary, for example, with adult cells, phytohemagglutinin, a chemical that stimulates growth and division is added. Then, after growth, colcemid, a chemical that stops the cells in mitosis is added. Then these cells are swollen in a low-salt solution and dropped onto a microscope slide. This step bursts open the cells and spreads out the chromosomes. The chromosomes are then stained and photographed through the microscope. In the past, photographs of chromosomes were literally cut up with scissors and arranged by size and shape. Now, this is all done with image-analysis software.

pairs must be faithfully copied. Given this huge number, even with an extremely accurate DNA copying process errors are still bound to occur. What type of errors can occur and what are their effects?

One type of error is the substitution of one base for another during copying. There are many different possible outcomes from such an error. For example, if the gene sequence is

CTT TGC AGT GCC CTC CAG AAA ATA AAG TAA

Figure 7.4 Chromosome Spread of an XXY Human. This chromosome spread was made using a special technique that highlights the sex chromosomes, making them darker than the other twenty-two pairs of chromosomes. The Y chromosome is the dark spot towards the top left, the two dark larger chromosomes are the X chromosomes. Photo courtesy of Norah McCabe.

Figure 7.4 Chromosome Spread of an XXY Human. This chromosome spread was made using a special technique that highlights the sex chromosomes, making them darker than the other twenty-two pairs of chromosomes. The Y chromosome is the dark spot towards the top left, the two dark larger chromosomes are the X chromosomes. Photo courtesy of Norah McCabe.

it codes for a protein with the following amino acid sequence (see figure 4.6):

If a DNA replication error results in G (in bold) at the third nucleotide of the first codon rather than T, which is the correct nucleotide,

CTG TGC AGT GCC CTC CAG AAA ATA AAG TAA

there will be no difference in the resulting amino acid because both CTT and CTG code for the same amino acid, leucine (see the genetic code in figure 4.6). This change is called a base substitution, but here there is no amino acid substitution.

On the other hand, if an error results in the following substitution at the first nucleotide position,

ATT TGC AGT GCC CTC CAG AAA ATA AAG TAA

then isoleucine, instead of leucine, will be inserted in the growing protein. Therefore, this base substitution results in an amino acid substitution. But, because leucine and isoleucine are very similar chemically, in many cases there will be no detectable difference in the protein function and thus no change in phenotype.

But, if the error results in a substitution at the second nucleotide position of the same codon,

CGT TGC AGT GCC CTC CAG AAA ATA AAG TAA

then arginine will be substituted for the leucine. These two amino acids are very different chemically. So, depending on the part of the protein that is affected, this error may result in a major change in function or make the protein nonfunctional.

Finally, there may be an error such as the following:

CTT TGC AGT GCC CTC TAG AAA ATA AAG TAA

Since TAG is one of three stop codons, the protein will end there rather than at the original stop codon TAA. Often such an error results in a shortened, nonfunctional protein. However, if the error occurs near the normal end of a large protein, as in this case, it may still result in the making of a partially functional protein. Therefore, the effect of changes in the base pairs of a gene can range from no effect at all to causing a major alteration that results in a totally nonfunctional protein.

The base sequence of DNA can be modified in other ways. Occasionally, an additional base may be inserted within a normal sequence. Alternatively, one of the bases may be skipped. These are called insertions and deletions, respectively. Let us look at the effect of the insertion of a base in the second codon of the following sequence

CTT TGC AGT GCC CTC CAG AAA ATA AAG TAA

which codes for the following sequence of amino acids.

Let us now insert an extra T (in bold) after the third T in our original sequence. We then get

CTT TTG CAG TGC CCT CCA GAA AAT AAA GTA A

this sequence now codes for a protein with the following amino acid sequence.

Note that from the point of insertion on, all the amino acids are different from the original ones! But more devastating than this, though this is a major problem in and of itself, now there is no stop codon in this sequence anymore! Thus this single base insertion results in a much longer protein than normal, with totally different amino acids! Similar situations occur in the case of a deletion. Try deleting one of the bases near the beginning of the first sequence above. Then use the genetic code in figure 4.6 to determine the amino acids of the mutated protein. Does your deletion mutation also make a larger protein by eliminating the stop codon?

An example of a DNA base change that results in a major defect is Marfan syndrome, a dominant trait discussed in chapter 3. Numerous base additions or deletions in a specific gene result in Marfan syndrome. The gene that is defective in Marfan syndrome, fibrillin, is very large, and most of that gene is important for its proper function. Because of its large size, mistakes can occur over a long sequence of DNA; that is, the gene presents a large target for mutations. Thus many defects in Marfan syndrome are new mutations rather than ones inherited by affected individuals. One example is a CGC to CCC change that results in a change from the amino acid arginine to the amino acid proline. There are other examples of single amino acid substitutions in the large protein that causes Marfan syndrome.

Amino acid substitutions can also affect the function of hemoglobin, the protein responsible for carrying oxygen in our blood. As we have already seen, sickle-cell anemia is caused by glutamic acid, present in normal hemoglobin at the sixth position, being changed to va-line. However, the amino acid right next to that normal glutamic acid is again glutamic acid in the seventh position. Variants have been found at that position, where either AAG, which codes for lysine, or GGG, which codes for glycine, replaces the normal GAG-coded glu-tamic acid. Interestingly enough, these hemoglobins function normally, unlike sickle-cell hemoglobin. Farther down the hemoglobin protein, in position 145, the normal amino acid is tyrosine. In two different mutations, either histidine or aspartic acid replaces this amino acid. Tyrosine, histidine, and aspartic acid are very different chemically, but instead of causing a defective protein, one might say that these mutations improve it, because these mutated hemoglobins have higher affinity for oxygen. These mutations have been found in individuals living at higher altitude, where higher affinity for oxygen improves survival. Table 7.1 shows some other single amino acid changes in hemoglobin and their effects.

Thus, a change in DNA sequence can result in a change of amino acid, but the effect of this change can be dramatically different depending upon the position of the amino acid and the amino acid that replaces it. Some changes are totally unnoticeable, while others cause devastating diseases. Others yet actually "improve" the function of the protein.

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