Weaver/Molecular Biology

Directly Undoing DNA Damage

One way to cope with DNA damage is to repair it, or restore it to its original, undamaged state. There are two basic ways to do this: (1) Directly undo the damage, or (2) remove the damaged section of DNA and fill it in with new, undamaged DNA. Let us begin by looking at two methods E. coli cells use to directly undo DNA damage.

In the late 1940s, Albert Kelner was trying to measure the effect of temperature on repair of ultraviolet damage to DNA in the bacterium Streptomyces. However, he noticed that damage was repaired much faster in some bacterial spores than in others kept at the same temperature. Obviously, some factor other than temperature was operating. Finally, Kelner noticed that the spores whose damage was repaired fastest were the ones kept most directly exposed to light from a laboratory window. When he performed control experiments with spores kept in the dark, he could detect no repair at all. Renato Dulbecco soon observed the same effect in bacteria infected with UV radiation-damaged phages. It now appears that most forms of life share this important mechanism of repair, which is termed photoreactivation, or light repair. However, placental mammals, including humans, do not have a photoreactivation pathway.

It was discovered in the late 1950s that photoreactivation is catalyzed by an enzyme called photoreactivating enzyme or DNA photolyase. This enzyme operates by the mechanism sketched in Figure 4. First, the enzyme detects and binds to the damaged DNA site (a pyrimidine dimer). Then the enzyme absorbs visible light, which activates it so it can break the bonds holding the pyrimidine dimer together. This restores the pyrimidines to their original independent state. Finally, the enzyme dissociates from the DNA; the damage is repaired.

Organisms ranging from E. coli to human beings can directly reverse another kind of damage, alkylation of the O6 of guanine. After DNA is methylated or ethylated, an enzyme called O6 methylguanine methyl transferase comes on the scene to repair the damage. It does this by accepting the methyl or ethyl group itself, as outlined in Figure 5.

The acceptor site on the enzyme for the alkyl group is the sulfur atom of a cysteine residue. Strictly speaking, this means that the methyl transferase does not fulfill one part of the definition of an enzyme-that it be regenerated unchanged after the reaction. Instead, this protein seems to be irreversibly inactivated, so we call it a "suicide enzyme" to denote the fact that it "dies" in performing its function. The repair process is therefore expensive; each repair event costs one protein molecule.

One more property of the O6-methylguanine methyl transferase is worth noting. The enzyme, at least in E. coli, is induced by DNA alkylation. This means bacterial cells that have already been exposed to alkylating agents are more resistant to DNA damage than cells that have just been exposed to such mutagens for the first time.

Summary: Ultraviolet radiation damage to DNA (pyrimidine dimers) can be directly repaired by a DNA photolyase that uses energy from visible light to break the bonds holding the two pyrimidines together. O6-alkylations on guanine residues can be directly reversed by the suicide enzyme O6-methylguanine methyl transferase, which accepts the alkyl group onto one of its amino acids.

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