Structure of p15(PAF)-PCNA complex and implications for clamp sliding during DNA replication and repair

The intrinsically disordered protein p15(PAF) regulates DNA replication and repair by binding to the proliferating cell nuclear antigen (PCNA) sliding clamp. We present the structure of the human p15(PAF)-PCNA complex. Crystallography and NMR show the central PCNA-interacting protein motif (PIP-box) of p15(PAF) tightly bound to the front-face of PCNA. In contrast to other PCNA-interacting proteins, p15(PAF) also contacts the inside of, and passes through, the PCNA ring. The disordered p15(PAF) termini emerge at opposite faces of the ring, but remain protected from 20S proteasomal degradation. Both free and PCNA-bound p15(PAF) binds DNA mainly through its histone-like N-terminal tail, while PCNA does not, and a model of the ternary complex with DNA inside the PCNA ring is consistent with electron micrographs. We propose that p15(PAF) acts as a flexible drag that regulates PCNA sliding along the DNA and facilitates the switch from replicative to translesion synthesis polymerase binding.We thank P. Redondo for help with the DNA duplex formation, T. Kaminishi for help with molecular modelling, M. Lelli for help with NMR relaxation measurements, S. Pongor for help with fluorescence polarization, N. Mailand for pointing out the p15 sequence similarity with histone H3 and S. Onesti for helpful discussions. This work was supported by the Spanish MINECO grants CTQ2011-28680 to F.J.B., BFU2011-23815 to G.M., and Juan de la Cierva to A. De. B., grant NNF14CC0001 to G.M., and SPIN-HD-ANR Chaires d'Excellence and the ATIP-Avenir to P. B. This research received funding from the European Community's FP7 grant agreement number 283570 (BioStruct-X, providing acces to SLS and ALBA syncrotrons) and 261863 (Bio-NMR, providing access to the 1 GHz NMR spectrometer at the Rhone-Alpes European Large Scale Facility for NMR).S