Buffer frames were then subtracted using PRIMUS. From the SAXS data, a radius of gyration (Rg) of 3.23 nm was calculated, which describes the average particle dimension in solution. The ab initio model fit from DAMMIF shows a χ 2 of 1.127, indicating good agreement with the experimental data (Figure and SI Table ). The corresponding dimeric envelope is highlighted in Figure , superimposed with the calculated dimeric model of Hsp90α CTD. Further, we tested the effect of 5b on the Hsp90 CTD dimer using SAXS (Figure ). Due to the low solubility of 5b, we needed to measure the sample as an ensemble of species in solution. First, we tested Hsp90α CTD without 5b on Xeuss 2.0 with Q-Xoom system and observed an increase of Rg to 3.40 nm. This increase is likely due to the small amount of tetramer in solution in the ensemble. Theoretically, the Rg of a tetrameric Hsp90α CTD species is 3.78 nm, using CRYSOL, indicating that even in batch mode SAXS measurements, the Hsp90α CTD protein is predominantly in a dimeric state. We added 5b with an equimolar concentration to Hsp90α CTD protein, and the Rg value slightly increased from 3.40 nm to 3.46 nm. However, with an increasing concentration of 5b to 1 mM, the Rg value increased to 4.11 nm. Compared to the theoretical Rg value of 3.78 nm for the tetramer, we observed that 5b induces oligomerization of Hsp90α CTD to species even larger than the tetrameric form (Figure ). It is unclear, however, if the oligomers are formed from Hsp90α CTD monomers or dimers, as the single species could not be resolved in the ensemble measurement.