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FIGURE 9-19 Crystal structure of a translesion polymerase. Shown here is the structure of a translesion (Y-iamtly DNA) poly merase, m gray, it) complex with template DMA, in purple, and an incoming nucleotide, in red (Ling H, Boudsoccj F„ Woodgate R, and Yang W. 2001. Cell 107: 9Î 102. Image prepared with Rob-Script, MolScript, and Raster 3D.)

FIGURE 9-19 Crystal structure of a translesion polymerase. Shown here is the structure of a translesion (Y-iamtly DNA) poly merase, m gray, it) complex with template DMA, in purple, and an incoming nucleotide, in red (Ling H, Boudsoccj F„ Woodgate R, and Yang W. 2001. Cell 107: 9Î 102. Image prepared with Rob-Script, MolScript, and Raster 3D.)

An important feature of these polymerases is that, although they are template dependent, they incorporate nucleotides in a manner that is independent of base pairing. This explains how the enzymes can synthesize DNA over a lesion on the template strand. Rut, because the enzyme is not "reading" sequence information from the template, iranslesion synthesis is often highly error-prone. Consider the case of an apurtnic or apyrmidinic site in which the lesion contains no base-specific information. The translesion polymerase synthesizes across triO lesion by inserting nucleotides in a manner that is not guided by base pairing. Nonetheless, the nucleotide incorporated may not be random—some translesion polymerases incorporate specific nucleotides. For example, a human member of the Y-family of translesion polymerases correctly inserts two A residues opposite a thymine dimer.

Box 9-3 The Y-Family of DNA Polymerases

DMA polymerases can be grouped into families, shown in various colors in the figure, based on their amino acid sequence similarities to each other. Recently, UmuC and certain other translesion DNA polymerases have been discovered to be founding members of a large and distinct family of DNA polymerases known as the Y-family, which are found in all three domains of life, Bacteria, Ardiaea, and Eukary ota. Members of the Y-famiiy of DNA polymerases charactenstically carry out DMA synthesis with kiw fidelity on undamaged DNA Templates but have the capacity to bypass lesions in DNA that block replication by members of the other families of DMA polymerases. Box 9-3 Figure 1 shows a phylogenefic tree for the Yfamiiy of translesion DMA polymerases.

Mtu DinP

Lla UmuC

Eta pADl UvrA

Psy RulB Sth R27 tvkicB

BOX 9-3 FIGURE 1 the phylogenefic tree of the Y-family of DNA polymerases. (Source: Adapted from Ohmon H. et at. Letter to the editor: Ihe Y-iamity of DNA polymerases. Mo! Cell 8; 7, fig 1.)

Mtu DinP

Lla DinP Mm Dinbl

HSOINB1 AIDInBh Ce OinSh Sp DinBh

Ce REVlh At REVIh ScR£V1 Sp REV1 Mm R£Vfh Ha REV1 Dm REV1h

Mm RadMB Hs

Bha BH1472 BstiYqjH Bfia BB2741 BsuYqjW

Ban pOX2 69 Bsu SPBC2 UvrX Spn QrfT3

Lla UmuC

Eta pADl UvrA

Ce Rad30h Hs RatOOA Mm Rad30A Dm Rat)30A R«j030h Lm UmuC

Psy RulB Sth R27 tvkicB

Eco R391 RumB Eco UmuC Sty UmuC Sty SamB StyTP110 ImpB Eco pKM101 WlucB Sma MucB

BOX 9-3 FIGURE 1 the phylogenefic tree of the Y-family of DNA polymerases. (Source: Adapted from Ohmon H. et at. Letter to the editor: Ihe Y-iamity of DNA polymerases. Mo! Cell 8; 7, fig 1.)

Because of its high error rate, translesion synthesis can be considered a system of last resort. It enables the cell to survive what might otherwise be a catastrophic block to replication but the price that is paid is a higher level of mutagenesis, for this reason, in E. coli the translesion polymerase is not present under normal circumstances. Rather, its synthesis is induced onty in response to DNA damage. Thus, the genes encoding the translesion polymerase are expressed as pari of a pathway known as the SOS response. Damage leads to the proteolytic destruction of a transcriptional repressor (the LexA repressor) which controls expression of genes involved in the SOS response including those for UrnuC and UmuD, the inactive precursor for UnmD'. Interestingly, the same pathway is also responsible for the proteolytic conversion of UmuD to UmuD'. Cleavage of LexA and UmuD are bodi stimulated by a protein called KecA, which is activated by single-stranded DNA resulting from DNA damage. RecA is a dual-function protein that is also involved in DNA recombination as we shall see in Chapter 10.

Finally, trans lesion synthesis* puses several fascinating and as yet unanswered questions. How does the translesion polymerase recognize a stalled replication fork? How does the translesion enzyme replace the normal implicative polymerase in the DNA replication complex? Once DNA synthesis is extended across the lesion, how does the norma! replicative polymerase switch back to and replace the translesion enzyme at the replication fork? Thmslesion polymerases have low processivity, so perhaps they simply dissociate bom the template shortly after copying across a lesion. Nonetheless, this explanation still leaves us with the challenge of understanding how the normal processive Enzyme is able to reenter the replication machinery.

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