A Slow ATP-induced Conformational Change Limits the Rate of DNA Binding but Not the Rate of β Clamp Binding by the Escherichia coli γ Complex Clamp Loader

摘要: In Escherichia coli, the γ complex clamp loader loads β-sliding onto DNA. The β tethers DNA polymerase III to and enhances efficiency of replication by increasing processivity synthesis. presence ATP, binds form a ternary complex. Binding primed template triggers hydrolyze ATP release Here, we investigated kinetics forming measuring rates binding A fluorescence intensity-based assay was developed in which pyrene covalently attached increases when bound Using this assay, an association rate constant 2.3 × 107 m−1 s−1 for determined. same experiments preincubated with before adding or added directly ATP. contrast, is faster than at time. Slow absence preincubation result rate-limiting ATP-induced conformational change. Our results strongly suggest that changes promote differ. slow change precedes may provide kinetic preference bind during loading reaction cycle.