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. 2020 Feb 3;30(3):381–395.e8. doi: 10.1016/j.cub.2019.11.058

Figure 6.

Figure 6

SA Binds to the A Subunit of PP2A

(A) DARTS assay suggests that PP2AA1 is potential target of SA. pPP2AA1::PP2AA1-GFP seedlings were used for the protein isolation. Samples were treated with DMSO (mock) and SA and digested by different concentrations of Pronase. Samples were further analyzed by western blot with an anti-GFP antibody.

(B) DSC analysis suggesting the potential binding of SA to recombinant His-PP2AA1. 5 μM of purified His-PP2AA1 protein was analyzed by DSC with or without 50 μM SA. Tm = 48.01°C and 45.03°C for His-PP2AA1+DMSO and His-PP2AA1+SA, respectively.

(C) SPR analysis of the His-PP2AA1 and SA interaction. An active synthetic SA analog (SA-f) was immobilized on a CM-5 sensor chip, and different concentrations of His-PP2AA1 were applied. The binding curve was plotted by values at the steady state, for which the sensorgram is shown is Figure S7D. A KD value of 3.623 μM was detected.

(D) SPR assay reveals the binding of His-PP2AA3 to SA. The same sensor chip as above was used, and different concentrations of His-PP2AA3 were applied. The binding curve was plotted by values at the steady state, with the data points shown in the sensorgram in Figure S7E. A KD value of 1.916 μM was detected.

See also Figures S5, S6, and S7.