Structure-based predictions of Rad1, Rad9, Hus1 and Rad17 participation insliding clamp and clamp-loading complexes

The repair of damaged DNA is coupled to the completion of DNA replication b y several cell cycle checkpoint proteins, including, for example, in fissio n yeast Rad1(Sp), HUs1S(p), Rad9(Sp) and Rad17(Sp), We have found that thes e four proteins are conserved with protein sequences throughout eukaryotic evolution, Using computational techniques, including fold recognition, comp arative modeling and generalized sequence profiles, we have made high confi dence structure predictions for the each of the Rad1, Hus1 and Rad9 protein families (Rad17(Sc), Mec3(Sc) and Ddc1(Sc) in budding yeast, respectively) . Each of these families was found to share a common protein fold with that of PCNA, the sliding damp protein that tethers DNA polymerase to its templ ate. We used previously reported genetic and biochemical data for these pro teins from yeast and human cells to predict a heterotrimeric PCNA-like ring structure for the functional Rad1/Rad9/Hus1 complex and to determine their exact order within it. In addition, for each individual protein family, co ntact regions with neighbors within the PCNA-like ring were identified, Bas ed on a molecular model for Rad17(Sp), we concluded that members of this fa mily, similar to the subunits of the RFC clamp-loading complex, are capable of coupling ATP binding with conformational changes required to load a sli ding clamp onto DNA, This model substantiates previous findings regarding t he behavior of Rad17 family proteins upon DNA damage and within the RFC com plex of clamp-loading proteins.