PRODUCTION, PURIFICATION, AND CRYSTALLIZATION OF HUMAN INTERLEUKIN-1-BETA CONVERTING-ENZYME DERIVED FROM AN ESCHERICHIA-COLI EXPRESSION SYSTEM

Interleukin-1 beta converting enzyme (ICE) is a cysteine protease that catalyzes the conversion of the inactive precursor form of IL-1 beta to an active mature form. The mature form of IL-1 beta is involved in mediating inflammatory responses and in the progression of autoimmune diseases. We recently reported on the production of active human ICE i n insect cells using the baculovirus expression system (Wang XM et al. , 1994, Gene 145:273-277). Because the levels of expression achieved w ith this system were limiting for the purpose of performing detailed b iochemical and biophysical studies, we examined the production of ICE in Escherichia coli. By using a tac promoter-based expression system a nd fusion to thioredoxin we were able to recover high levels of active ICE protein. The expressed protein, which was distributed between the soluble and insoluble fractions, was purified to homogeneity from bot h fractions using a combination of classical and affinity chromatograp hy. Comparisons of ICE derived from both fractions indicated that they were comparable in their specific activities, subunit composition, an d sensitivities to specific ICE inhibitors. The combined yields of ICE obtained from the soluble and insoluble fractions was close to 1 mg/L of induced culture. Recombinant human ICE was crystallized in the pre sence of a specific ICE inhibitor in a form suitable for X-ray crystal lographic analysis. This readily available source of ICE will facilita te the further characterization of this novel and important protease.