DNA Repair Mechanisms Part-02

# DNA Repair Mechanisms:

Damage to DNA caused by replication errors or mutations may have serious consequences. The multiplicity of repair mechanisms that have evolved in organisms ranging from bacteria to humans emphatically documents the importance of keeping mutation at a tolerable level. Some types of DNA repair mechanisms are –

1)      light-dependent repair or photoreactivation

2)       excision repair

                     I.            Base Excision Repair

                   II.            Nucleotide Excision Repair

3)      Mismatch repair

4)      postreplication repair

5)      the error-prone repair system (SOS response).

6)      Double-strand break repair.

# 1.Light-Dependent Repair:

Light-dependent repair or photoreactivation of DNA in bacteria is carried out by a light activated enzyme called DNA photolyase. When DNA is exposed to ultraviolet light, thymine dimers are produced by covalent cross-linkages between adjacent thymine residues (see Figure 13.12b)

DNA photolyase recognizes and binds to thymine dimers in DNA, and uses light energy to cleave the covalent cross-links (Figure 13.25). Photolyase will bind to thymine dimers in DNA in the dark, but it cannot catalyze cleavage of the bonds joining the thymine moieties without energy derived from visible light, specifically light within the blue region of the spectrum. Photolyase also splits cytosine dimers and cytosine-thymine dimers. Thus, when ultraviolet light is used to induce mutations in bacteria, the irradiated cells are grown in the dark for a few generations to maximize the mutation frequency.

# 2.Excision Repair:

Excision repair of damaged DNA involves at least three steps.

v  In step 1, a DNA repair endonuclease or endonuclease-containing enzyme complex recognizes, binds to, and excises the damaged base or bases in DNA.

v  In step 2, a DNA polymerase fills in the gap by using the undamaged complementary strand of DNA as template.

v  In step 3, the enzyme DNA ligase seals the break left by DNA polymerase to complete the repair process.

There are two major types of excision repair:

a)       base excision repair systems remove abnormal or chemically modified bases from DNA,

b)      nucleotide excision repair; pathways remove larger defects like thymine dimers.

Both excision pathways are operative in the dark, and both occur by very similar mechanisms in E. coli and humans.

# 2(a). Base Excision Repair Mechansim:

Base excision repair (Figure 13.26) can be initiated by any of a group of enzymes called DNA glycosylases that recognize abnormal bases in DNA.

·         Each glycosylase recognizes a specific type of altered base, such as deaminated bases, oxidized bases, and so on (step 2).

·         The glycosylases cleave the glycosidic bond between the abnormal base and 2-deoxyribose, creating apurinic or apyrimidinic sites (AP sites) with missing bases (step 3).

·         AP sites are recognized by enzymes called AP endonucleases, which act together with phosphodiesterases to excise the sugar-phosphate groups at these sites (step 4).

·         DNA polymerase then replaces the missing nucleotide according to the specifications of the complementary strand (step 5),

·         and DNA ligase seals the nick (step 6).

# 2(b). Nucleotide Excision Repair Mechanism:

Nucleotide excision repair removes larger lesions like thymine dimers and bases with bulky side-groups from DNA.

In nucleotide excision repair, a unique excision nuclease activity produces cuts on either side of the damaged nucleotide(s) and excises an oligonucleotide containing the damaged base(s). This nuclease is called an excinuclease to distinguish it from the endonucleases and exonucleases that play other roles in DNA metabolism.

The E. coli nucleotide excision repair pathway is shown in Figure 13.27. In E. coli, excinuclease activity requires the products of three genes, uvrA, uvrB, and uvrC (designated uvr for UV repair). A trimeric protein containing two UvrA polypeptides and one UvrB polypeptide recognizes the defect in DNA, binds to it, and uses energy from ATP to bend the DNA at the damaged site.

The UvrA dimer is then released, and the UvrC protein binds to the UvrB/DNA complex. The UvrC protein cleaves the fourth or fifth phosphodiester bond from the damaged nucleotide(s) on the 3´ side and the eighth phosphodiester linkage from the damage on the 5´ side. The uvrD gene product, DNA helicase II, releases the excised dodecamer.

In the last two steps of the pathway, DNA polymerase I fills in the gap, and DNA ligase seals the remaining nick in the DNA molecule.

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