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. Author manuscript; available in PMC: 2015 Nov 10.
Published in final edited form as: Science. 2015 Oct 8;350(6259):404–409. doi: 10.1126/science.aac5789

Fig. 1. Preparation and characterization of NAIP-NLRC4Δ complexes.

Fig. 1

(A) Domain organizations of Salmonella typhimurium PrgJ, mouse NAIP2, mouse NLRC4, and mouse caspase-1. Domain size is drawn approximately to scale; residue numbers are labeled. (B) SDS–polyacrylamide gel electrophoresis (PAGE) of different fractions of the sucrose gradient ultracentrifugation during the purification of the PrgJ-NAIP2-NLRC4Δ complex. Locations of the three component proteins are labeled. The asterisk indicates a contaminating band. (C)A representative negative-stain EM image from fraction 7 in (B). (D) SDS-PAGE of amylose resin elution (lane 1), anti-Flag flow-through (lane 2), and anti-Flag elution (lane 3) fractions during the purification of the coexpressed His-FliC– Flag-NAIP5–His-MBP-NLRC4Δ complex. An enlarged image of lane 3 is shown. (E) Ni-NTA gold labeling (5 nm) of purified His-Sumo-PrgJ–NAIP2ΔBIR-His– His-Sumo-NLRC4Δ complex upon removal of the His-Sumo tag. (F) Schematic diagram of partial and complete inflammasome particles that contain variable ratios between NAIP2 (yellow) and NLRC4 (cyan). (G) Representative cryo-EM micrograph of PrgJ-NAIP2-NLRC4Δ particles. (H) An averaged 2D class of the 11-bladed PrgJ-NAIP2-NLRC4Δ inflammasome complex. The dimensions of the image are 43.5 nm × 43.5 nm.