Figure 2. The affinity of p60 for microtubules is unchanged by a phosphomimetic mutation.

250 nM X. laevis p60 or p60^S131E with either ATP (a) or ADP (b) was added to increasing concentrations of taxol-stabilized MTs from 0-10 μM in BRB80 with 10μM Taxol. Reactions were incubated at room temperature for 15 min, pelleted by centifugation and analyzed by SDS-PAGE. Binding data was fit to the quadratic binding equation as previously described²⁰. Error in fitted parameters was estimated by monte carlo sampling within the experimentally determined error and fitting the resulting datasets²⁷. This procedure generates hundreds of theoretical datasets that fit the measured data by randomly sampling a single value at each protein concentration that falls within the experimental error of that data point. The theoretical datasets are individually fitted normally. The maximum likelihood mean±SD was determined from 250 individual fits and provides the reported K_D value and error. Plotted data is from two independent experiments with two separate preparations of each protein.