Crystal Structure Of PCNA-SIVA1(80-96) Complex

Tanslesion DNA synthesis (TLS) is a ubiquitous DNA damage tolerance mechanism among yeast and mammals. It is capable of bypassing DNA lesions that tend to block replication by recruiting a special class of TLS polymerases for repair at a later time point. A key event in this regulation is the monoubiquitination of proliferating cell nuclear antigen (PCNA) by RAD6-RAD18 ubiquitin-conjugating/ligase complex. It is reported that this process is mediated by SIVA1 serving as a molecular bridge between Radl8 and PCNA. However, the detailed structural mechanism concerning this process remains largely unknown. In this study, we particularly investigated the interaction between PCNA and the PIP-motif-containing peptide derived from SIVA1 by using X-ray crystallography and isothermal titration calorimetry (ITC). Our crystallographic and thermodynamic data show that SFVA1 (80-90) peptide tightly binds to the front face of PCNA near the IDCL (the interdomain connector loop) region with a 1:1 stoichiometry and a dissociation constant (Kp) in a micromolar range, which resemble to those of many other PIP-motif-containing peptides. Our results not only provide the direct evidence of the interection between PCNA and SIVA1, but also shed some lights on the molecular basis of the regulation of TLS by SIVA1.