## 6 Global Genome Repair (GGR)
### Dissociation of UvrA from DNA and Formation of the Pre-Incision Complex  
The dissociation of UvrA from the damage-sensing complex, the orchestration of the initial loading of UvrB and subsequent translocation to the lesion using its 5\({}^{\prime}\)\(\rightarrow\)3\({}^{\prime}\) ssDNA translocation activity, the number of UvrB molecules loaded by UvrA and the identity of the DNA strand presented to UvrC have been enduring mysteries of the NER pathway. It has long been known that dissociation of UvrA from the damage sensing complex is a pre-requisite for loading of UvrB at the lesion to form the pre-incision complex [100], however, the mechanisms that underlie this process are incompletely understood. Prior studies have established that ATP hydrolysis by UvrA leads to its dissociation from the DNA [115] and that ATP binding and hydrolysis accompany translocation of UvrB towards the lesion [53; 54]. The possibility that UvrA could load two UvrB molecules on distinct strands, approximately 80 A away on either side of the lesion [21], provides a structural framework for understanding how the UvrA\({}_{2}\)B\({}_{2}\)*damaged DNA complex transitions to the UvrB*lesion pre-incision complex. This transition has been envisioned via two models:  
Figure 3: Two models for the UvrA\({}_{2}\)B\({}_{2}\) complex. The two UvrA protomers are shown in cyan and orange, while the two UvrB molecules are shown in different shades of gray. The crystal structure of the UvrA\({}_{2}\)B\({}_{2}\) complex has the UvrA dimer in the ‘closed-grooved’ configuration [22] **(left)**, whereas the negative stained electron microscopy structure of UvrA\({}_{2}\)B\({}_{2}\) complex in the presence of DNA has the UvrA dimer in the ‘open-tray’ configuration **(right)**[21]. Although dsDNA was included in the EM sample, the DNA is not visible in the electron density map and is not included in the model. The two UvrA\({}_{2}\)B\({}_{2}\) models are aligned on the orange UvrA protomer. With this alignment the cyan UvrA protomers in the two models are related by a 50\({}^{\circ}\) helical rotation (50\({}^{\circ}\) rotation, 20 Å translation) about the axis approximately parallel to the presumed DNA binding path. the conformational change and the translocation models [21]. In the first model, a large conformational change in the UvrA\({}_{2}\)B\({}_{2}\) complex delivers UvrB from its peripheral location in the complex to its final location at the lesion site; precise positioning at the lesion is enabled by helicase activity of UvrB, which accompanies dissociation of UvrA. By contrast, the translocation model implies that UvrA dissociates prior to translocation of UvrB to the site of the lesion [20,21]. In either model, loading two UvrB molecules on distinct strands could serve as a mechanism to identify the lesion-containing strand for presentation to the UvrC nuclease. The crystal structure of the UvrA\({}_{2}\)B\({}_{2}\) complex provided a structural basis for understanding how ATP hydrolysis at the UvrA proximal site and conformation switching by the signature domain II and possibly the UvrB-binding domain, enables dissociation [21]. A recent single molecule live cell imaging study showed that in the absence of UvrB, UvrA displays a lifetime of \(\sim\)20-30 s on damaged DNA; a short-lived complex (\(\sim\)1.4 s) was also observed but not extensively analyzed. The presence of UvrB reduced the lifetime of the long-lived population to \(\sim\)8.7 s, whereas the lifetime of the short-lived population remained unchanged. These measurements capture the dynamics of UvrA loading UvrB onto lesion DNA prior to its dissociation [27]. Notably, a prior single molecule study found no evidence that UvrB reduces the lifetime of UvrA on DNA in vitro [98]. The finding that UvrB may approach the lesion from the 5' side [116] has been more precisely defined by recent structural, crosslinking and biochemical studies. Two orthogonal experiments directly documented translocation. First, [PERSON] et al.