FIGURE 2.

Fluorescence polarization confirms that Hsp90-PP5 has higher affinity than Hsp70-PP5. A, raw mP values were plotted with control peptide. No binding was observed with increasing concentrations of PP5 and control peptide at 20 nm. B, direct binding was measured with increasing concentration of PP5 and 5-mer peptides (20 nm). C, direct binding was measured using increasing concentrations of purified full-length PP5 with 5FAM 10-mer-labeled peptide (20 nm). D, to see how the residues Met and Ile contribute affinity to PP5, HSP70–90 peptides were synthesized so that HSP90 10-mer peptide contained an Ile instead of a Met and HSP70 10-mer peptide contained an Met instead of an Ile. Fluorescence polarization (FP) was performed with increasing concentrations of PP5, and both peptides were held constant at 20 nm. Data are presented as mean ± S.D. (n = 3).