Structure of Monoubiquitinated PCNA and Implications for Translesion Synthesis and DNA Polymerase Exchange

DNA synthesis by classical polymerases can be blocked by many lesions. These blocks are overcome by translesion synthesis, whereby the stalled classical, replicative polymerase is replaced by a non-classical polymerase. In eukaryotes, this polymerase exchange requires PCNA monoubiquitination. To better understand the polymerase exchange, we have developed a novel means of producing monoubiquitinated PCNA, by splitting the protein into two self-assembling polypeptides. We determined the X-ray crystal structure of monoubiquitinated PCNA and found that the ubiquitin moieties are located on the back face of PCNA and interact with it via their canonical hydrophobic surface. Moreover, the attachment of ubiquitin does not change PCNA’s conformation. We propose that PCNA ubiquitination facilitates non-classical polymerase recruitment to the back of PCNA by forming a new binding surface for non-classical polymerases, consistent with a “tool belt” model of the polymerase exchange.