Fig. 6: Proposed models of function of Pol κ in TLS.
From: Cryo-EM structure of human Pol κ bound to DNA and mono-ubiquitylated PCNA
a Pol κ interaction with mono-ubiquitylated PCNA. Pol κ binds PCNA encircling DNA through the internal PIP-box at the C-terminus of the PAD. If the PIP-box interaction is lost, Pol κ UBZ zinc fingers interact with the ubiquitin molecules flexibly attached to the back face of Ub-PCNA, helping retain the polymerase to the DNA primer/template junction. Ubiquitins are modeled as in the cryo-EM structure of Pol κ−DNA−Ub-PCNA complex (Ub1 position), but may occupy alternate positions due to their intrinsic flexibility. The orange dotted line represents the C-terminal disordered region connecting the Pol κ core to the UBZ domains. The homology model for Pol κ UBZ/Ubiquitin complex was built as shown in Fig. 5. b PCNA-directed polymerase swapping in TLS. At a lesion on the DNA template strand (highlighted in red), Pol δ holoenzyme stalls, and Pol κ is recruited to PCNA following two alternative paths. Either Pol δ dissociates into solution prior to or during Pol κ binding to PCNA and P/T DNA (sequential model), or Pol δ tilts remaining attached to the PIP-box site on PCNA and Pol κ is recruited to the exposed PIP-box site, capturing the P/T DNA released by Pol δ (toolbelt model). Tilting of Pol δ is achieved by disruption of two of the three indicated contact points with PCNA36, and allows to accommodate actively synthesizing Pol κ on PCNA in the “active state” without steric clashes.
