ATP bound to the origin recognition complex is important for preRC formation

The origin recognition complex (ORC) binds origins of replication and direc ts the assembly of a higher order protein complex at these sites. ORC binds and hydrolyzes ATP in vitro. ATP binding to the largest subunit of ORC, Or c1p, stimulates specific binding to origin DNA; however, the function of AT P hydrolysis by ORC is unknown. To address the role of ATP hydrolysis, we h ave generated mutants within Orc1p that are dominant lethal. At physiologic al ATP concentrations, these mutants are defective for ATP hydrolysis but n ot ATP binding in the absence of DNA, These mutants inhibit formation of th e prereplicative complex when overexpressed. The dominant lethal phenotype of these mutant ORC complexes is suppressed by simultaneous overexpression of wild-type, but not mutant, Cdc6p. Our findings suggest that these hydrol ysis-defective mutants inhibit growth by titrating Cdc6p away from the orig in. Based an these observations, we propose that Cdc6p specifically recogni zes the ATP-bound state of Orc1p and that ATP hydrolysis is coupled to preR C disassembly.