Mutagenesis (redirected from site directed mutagenesis)

mutagenesis

Genetics the generation, usually intentional, of mutations

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Mutagenesis 

the process that results in the appearance of permanent hereditary changes, or mutations. Mutagenesis can occur spontaneously or be induced by a variety of physical or chemical agents, called mutagens.

Underlying mutagenesis are changes in the structure of the nucleic acids, which store and transmit hereditary information. These changes are expressed as gene mutations or chromosomal aberrations. Disruption of the mitotic apparatus of cell division is also possible; such mutagenesis gives rise to genome mutations, such as polyploidy and aneuploidy. Damage to the nucleic acids, DNA and RNA, involves either rupture of the sugar and phosphate residues or insertion or deletion of nucleotides. Chemical changes can also occur in the nitrogenous bases, either directly reflecting gene mutations or leading to the appearance of such mutations during subsequent replication of the damaged nucleic acid molecule. There are a number of ways in which changes in the nitrogenous bases can occur. When a purine base is replaced by another purine base, or a pyrimidine base by another pyrimidine base, the mutation is called a transition. When a purine base is replaced by a pyrimidine base, or a pyrimidine base by a purine base, the resulting mutation is called a transversion. This leads to two types of disturbances in the nucleotide triplets, or codons, which determine protein synthesis: nonsense codons, which altogether fail to code for the inclusion of certain amino acids into the protein being synthesized, and missense codons, which signal the inclusion of an incorrect amino acid into the protein, thus altering the protein’s properties. The insertion or deletion of nucleotides causes a misreading of genetic information by shifting the encoding framework. This usually results in nonsense codons and, only rarely, in missense codons.

The various mutagens differ in the mechanism by which they induce mutagenesis. Ionizing radiation acts on the nucleic acids directly, ionizing and activating the atoms. This disrupts the sugar and phosphate residues of the nucleic acid molecule and the hydrogen bonds between the complementary DNA strands. It also produces cross-links between these strands and destroys the nitrogenous bases, especially pyrimidine. The direct action of ionizing radiation on the chromosomes and chromosomal DNA is responsible for the almost linear relationship between the radiation dose, on the one hand, and the frequency of induced gene mutations and minor deletions, on the other. However, the dose-frequency relationship is more complex in the case of those types of chromosomal aberrations that result from two chromosome breaks: larger deletions, inversions, and translocations, among others. The mutagenic action of ionizing radiation can be indirect: as the radiation passes through the cytoplasm or the nutrient medium in which the microorganisms are cultured, radiolysis of the water occurs, with the resulting appearance of free radicals and peroxides that subsequently act as the mutagens.

Ultraviolet radiation excites the electron shells of the atoms, which in nucleic acids triggers various chemical reactions that also can result in mutations. The most important of these reactions are the hydration of cytosine and formation of thymine dimers, but the rupture of the hydrogen bonds between the DNA strands and the formation of cross-links between these strands also play a part in mutagenesis. Ultraviolet rays do not readily penetrate tissues, and their mutagenic action is manifested only when they can reach the genetic apparatus, for example, during the irradiation of viruses, bacteria, and plant spores. Ultraviolet rays with a wavelength of 2500 angstroms (A) to 2800 A can be absorbed by the nucleic acids and are the most mutagenic. The rays in the visible spectrum suppress the mutagenic effect of ultraviolet radiation.

Alkylating compounds include the most powerful of the known mutagens (supermutagens), for example, nitroso-ethylurea and ethylmethanesulfonate. These compounds alkylate the phosphate radicals of the nucleic acids, which results in disturbances in the structure of the sugar and phosphate residues. Such compounds also alkylate the nitrogenous bases, especially guanine, resulting in an impairment of the precision of replication of the nucleic acid and the appearance of transitions and occasional transversions.

Analogs of nitrogenous bases can be incorporated into the nucleic acids, giving rise to transitions and transversions in subsequent replication. Nitrous acid also causes these types of changes, by deaminating the nitrogenous bases. Acridine dyes form a complex with DNA that interferes with the replication of the DNA. As a result, one or more pairs of nucleotides are lost or additionally inserted, causing the encoding framework to shift. Other chemical mutagens produce similar reactions with nucleic acids, but the mechanisms by which many of them effect mutagenesis are still obscure.

Mutagens can interfere with the cytoplasmic apparatus of mitosis. This results in nondisjunction of all the split chromosomes or in incorrect redistribution of the chromosomes among the daughter cells. Nondisjunction gives rise to polyploidy; incorrect redistribution gives rise to aneuploidy. Certain chemical agents, for example, the alkaloid colchicine, are known to act in this manner.

A variety of external factors may greatly influence the course of mutagenesis. For example, the frequency of mutations induced by ionizing radiation increases when there is a surplus of oxygen in the cell and decreases when there is a deficiency. Such a decrease can be achieved by carrying out irradiation in a nitrogen atmosphere. Some substances suppress mutagenesis. For example, the introduction of adenosine or guanosine into a cell inhibits the mutagenic action of analogs of purine bases, and the enzyme catalase diminishes the mutagenic effect of ionizing radiation.

After exposure to certain chemical mutagens, mutations may appear either immediately or some time later, occasionally after several cell generations.

REFERENCES

See references under .

S. M. GERSHENZON