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. 2013 Nov 12;8(11):e78443. doi: 10.1371/journal.pone.0078443

Figure 4. VER-155008 inhibits Hsp70's ATPase activity in a competitive manner and slows down nucleotide association.

Figure 4

Steady-state ATPase activity of Hsp70 (4 µM) in the absence (A, C) or presence (B, C) of 1 µM Hdj1. VER-155008 (A, B) and PES (C, D) were added as indicated. D, zoom of C for 0 to 12 µM ATP. KM and Ki values for VER-155008 were determined by global fitting of the data. Differences in KM and kcat in the presence of PES as compared to the DMSO control were not statistically significant (t-test, p values of 0.27 to 0.85). In contrast, differences in apparent KM observed in the presence of VER-155008 was statistically significant (p = 0.05 in absence of Hdj1; p = 0.005 in presence of Hdj1). Error bars represent the standard errors of at least three independent measurements. E, F, Association of MABA-ADP (0.5 µM) to nucleotide-free Hsp70 (0.5 µM) in the presence of VER-155008 (20 µM) and PES (160 µM). Changes in fluorescence intensity were followed in a stopped flow device for 20 (E) and 500 seconds (F). Comparison of the ADP association rates in the absence and presence of PES using the students t-test resulted in a P value of 0.41, indicating that the differences are not statistically significant.