DNA Repair and Recombiantion Methyldirected mismatch repair 1

DNA Repair and Recombiantion

Methyl-directed mismatch repair (1) • • • If any mismatch escapes the proof reading mechanisms it will cause distortion of the helix. This can be detected and repaired but it is important that the repair enzyme can distinguish the new strand from the old. This is possible in E. coli because there is an enzyme which methylates the A in a sequence GATC. This methylation does not occur immediately after synthesis and until it does the two strands are distinguishable.

Methyl-directed mismatch repair (2)

Repair of DNA damage in E. coli (1) • • • DNA damage occurs continually for a range of reasons including depurination, depyrimidation (to a lesser extent), ionizing radiation, free radicals, carcinogens, UV light. It is statistically unlikely for most of these changes, which affect one strand, that the second strand will be hit at the same time. This means the other strand can be used as template to repair the damaged strand.

Repair of DNA damage in E. coli (2) • • • UV light causes the formation of thymine dimer where adjacent Ts occur on the same strand. There is an enzyme which returns the thymines to their original form. Alkylation of bases can be reversed by the alkyl group being transferred to a protein, which is then destroyed. If the helix is distorted, for instance by T dimers, the distortion can be removed and replaced using the other strand as template.

Repair of double-strand breaks • • One mechanism involves the ligation of breaks. This is likely to change the sequence but it is statistically unlikely that the site will be critical. The second requires homologous recombination with the other chromosome in a diploid organism.

DNA damage repair in eukaryotes • • • The dealkylating proteins found in bacteria are also found in humans, as are enzymes repairing thymidine dimers and genes corresponding to Mut S and Mut L. Xeroderma pigmentosum is a genetic disease in which the capacity to repair thymine dimers is lost. People with this disease suffer skin cancer at high rates on exposure to sunlight. Mismatch repair proteins have also been found in humans. Mutations in these genes are a cause of colorectal cancer.

Homologous recombination • Homologous recombination is illustrated in Fig. 23. 25. • Recombination occurs naturally in cells. General recombination is the commonest type. In addition site specific recombination also occurs. • Homologous recombination allows reassortment of related but slightly different genes in an organism, thus generating genetic diversity. • This genetic diversity allows recombination of genes, generates genetic diversity and facilitates the operation of natural selection.

Mechanism of homologous recombination in E. coli • The process is best understood in this organism. • The first step requires single strand invasion, where a single strand inserts itself into the DNA duplex and displaces its homologous strand via a triple stranded intermediate. It is catalysed by the enzyme Rec. A. • A D loop is formed (Fig. 23. 26) and the process uses ATP.

Formation of cross-over junctions by single-stranded invasion • One strand of each DNA duplex is nicked to generate free 3´ ends. • These invade the homologous duplexes and result in displacement of the original partner. • The nicks are sealed to form Holliday junctions.

Figure 14. 05

Chi site: GCTGGTGG

Recombination in eukaryotes • Meiosis is accompanied by strand crossovers or chiasmata. Following DNA replication chromatids pair and recombination occurs. There are two rounds of cell division.