H1 Flagella Made

H2 rh1

repressor

H2 rh1

repressor

Ill flagella

Since many amino acids have several codons, changing the third base of a codon often leaves the amino acid unchanged.

is needed to guide base modifications to a neighboring exon (see Ch. 12). Nevertheless, most base changes within most introns are silent mutations.

The third main type of silent mutation occurs within the coding region of a gene and does get passed on to the messenger RNA. Remember that each codon, or group of three bases, is translated into a single amino acid in the final protein product. However, because there are 64 different codons, most of the 20 possible amino acids have more than one codon (see Codon Table, Fig. 8.02). So a base change that converts the original codon into another codon that codes for the same amino acid will have no effect on the final structure of the protein.

For example, the amino acid alanine has four codons: GCU, GCC, GCA and GCG. (Note that the sequences are discussed in RNA language; these are the codons as found on mRNA.) Since they all have GC as the first two bases, any codon of the form GCN (N = any base) will give alanine. A mutation in an original GCC sequence changing the last C to an A or a G or a U results in change in the sequence of the codon, but there is no change in the amino acid produced (alanine) in the resulting protein. Many other amino acids (such as valine, threonine and glycine) also have sets of four codons in which the last base does not matter. This pattern is referred to as third base redundancy. Very often, altering the third base of a codon has no effect on the protein that will be made. In other words, about a third of single base substitutions will be silent, third base redundancy Situation where a set of four codons all code for the same amino acid and thus the identity of the third codon base makes no difference during translation even if they occur within the protein coding region of a gene. [Note: the term "codon degeneracy" refers to the fact that a single amino acid may be encoded by multiple codons. In most, but not all cases, this is due to third base redundancy. However, both arginine and serine have six codons each; these inevitably differ among themselves by more than just the third base.]

Third base mutations that do not alter protein identity can sometimes have effects due to differential codon usage and tRNA bias. Different codons for the same amino acid often vary in their frequency of use and the corresponding tRNAs are often present in levels that are related to the frequency of codon usage. Consequently, if changing the third base converts a frequently used codon to a rarely used codon for the same amino acid, translation may be slowed due to shortage of the appropriate tRNA. (Conversely, changing a rare to a common codon may speed up translation.) This effect is usually only significant for proteins that are made at such high levels that their synthesis uses a significant proportion of the available tRNA in the cell.

DNA may be damaged by a variety of chemicals and by radiation.

Base analogs are mistaken by the cell for the natural nucleic acid bases.

Intercalating agents result in the insertion of an extra base pair during DNA replication.

Chemical Mutagens Damage DNA

Mutations that are caused by agents that damage the DNA are known as induced mutations. Agents that mutate DNA are called mutagens and are of three main types: mutagenic chemicals, radiation and heat. Even if there are no dangerous chemicals or radiation around, mutations still occur, though less frequently. These are spontaneous mutations. Some of these are due to errors in DNA replication. The enzymes of DNA replication are not perfect and sometimes make mistakes. In addition, DNA undergoes certain spontaneous chemical reactions (alterations) at a low but detectable rate and this rate goes up with increasing temperature.

The most common mutagens are toxic chemicals that react with DNA and alter the chemical structure of the bases. For example, EMS (ethyl methane sulfonate) is widely used by molecular biologists to mutagenize growing cells. It adds an ethyl group to bases in DNA and so changes their shape and their base-pairing properties. Nitrite converts amino groups to hydroxyl groups and so converts the base cytosine to uracil (Fig. 13.13). Nitrite is used experimentally to mutate purified DNA, such as a cloned gene carried on a plasmid, while the plasmid is in the test tube. The mutagenized DNA is then transferred back into a cell to identify the mutations that were generated. During DNA replication, the DNA polymerase misidentifies these altered bases and puts in the wrong bases in the new complementary strand of DNA it is making (Fig. 13.13).

Base analogs are chemical mutagens that mimic the bases found in natural DNA. For example, bromouracil resembles thymine in shape. It is converted by the cell to the DNA precursor, bromouridine triphosphate, which DNA polymerase inserts where thymine should go. Unfortunately, bromouracil can flip-flop between two alternative shapes (Fig. 13.14). In its alternate form, bromouracil resembles cytosine and pairs with guanine. If bromouracil is in its misleading form when DNA polymerase arrives, a G will be put into the new strand opposite the bromouracil instead of A.

Some mutagens imitate the structure of a base pair rather than a single base. For example, acridine orange has three rings and is about the size and shape of a base pair. Acridine orange is not chemically incorporated into the DNA. Instead, it squeezes in between the base pairs in the DNA (Fig. 13.15), a process referred to as intercalation. During DNA replication, the DNA polymerase mistakes the intercalating agent for a base pair and puts in an extra base when making the new strand. As discussed above, insertion of an extra base will change the reading frame of the protein encoded by a acridine orange A mutagenic agent that acts by intercalation base analog Chemical mutagen that mimics a DNA base induced mutation Mutation caused by external agents such as mutagenic chemicals or radiation intercalation Insertion of a flat chemical molecule between the bases of DNA, often leading to mutagenesis mutagen Any agent, including chemicals and radiation, that can cause mutations spontaneous mutation Mutation that occurs "naturally" without the help of mutagenic chemicals or radiation

Chemical Mutagens Damage DNA 349

A) Alkylating agents attack bases

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