Skip to main content
. 2009 Feb 17;106(10):3935–3940. doi: 10.1073/pnas.0809414106

Fig. 3.

Fig. 3.

Bcl-2 and Bcl-XL inhibit ATP binding to NLRP1. (A) His-6-NLRP1 (0.125 μM) was incubated for 15 min in ice in the presence of GST-Bcl-XL, His-Bcl-XLΔLoop, GST-Bcl-2, GST, or Bid (1 μM). The mixture was then incubated for additional 15 min in ice with 1 μM MDP-LD, FITC-conjugated ATP analog (10 nM) and Mg2+ (0.5 mM). ATP binding was analyzed by FPA (n = 3), measuring milliPolars (mP), and the percentage of inhibition was determined vs. NLRP1 incubated only with MDP-LD (mean ± SD). (B) His-6-NLRP1 (0.1 μM) was incubated for 15 min in ice with various amounts of GST-Bcl-2 (black squares), GST-Bcl-XL (white triangles), or GST-Bcl-XLΔLoop (black triangles) and then incubated 15 min on ice with 1 μM MDP-LD, FITC-conjugated ATP analog (10 nM) and Mg2+ (0.5 mM). ATP binding to NLRP1 was analyzed by FPA (n = 3), measuring milliPolars (mP), and the percentage of inhibition calculated (mean ± SD). Percentage inhibition values were derived by subtracting non-specific FITC-ATP binding, based on experiments where an excess of unlabeled ATP (100 nM) was applied to determine the competitive component of FITC-ATP binding (Fig. S2).