FIGURE 2.

The gp44/62 clamp loader complex binds four ATP molecules with similar affinities, both in the absence and in the presence of the gp45 clamp. The ATP bound to gp44/62 under steady-state conditions was measured (data points) using an ultrafiltration method as described under “Experimental Procedures.” Component concentrations were 30 μm gp44/62 alone (open symbols) or in the presence of 35 μm WT-gp45 (closed symbols). An ATP regeneration system was present in the reaction mixture. The curves were obtained by fitting the data points to a binding model (Equation 1) with four equal and independent ATP-binding sites.