Inhibition of interleukin-1 beta converting enzyme by the cowpox virus serpin CrmA. An example of cross-class inhibition.

Abstract

W e reported previously that human interleukin-lp converting enzyme (ICE) is regulated by the CrmA serpin encoded by cowpox virus.We now report the mechanism and kinetics of this unusual inhibition of a cysteine proteinase by a member of the serpin superfamily previously thought to inhibit serine proteinases only.CrmA possesses several characteristics typical of a number of inhibitory serpins.It is conformationally unstable, unfolding around 3 M urea, and stable to denaturation in 8 M urea upon complex formation with ICE.CrmA rapidly inhibits ICE with an association rate constant (ko,,) of 1.7 x lo' M -~ s-', forming a tight complex with an equilibrium constant for inhibition (ICi) of less than 4 x 10"' M.These data indicate that CrmA is a potent inhibitor of ICE, consistent with the dramatic effects of CrmA on modifying host responses to v i r u s infection.The inhibition of ICE by CrmA is an example of a "cross-class" interaction, in which a serpin inhibits a non-serine proteinase.Since CrmA possesses characteristics shared by inhibitors of serine proteinases, we presume that ICE, though it is a cysteine proteinase, has a substrate binding geometry strikingly close to that of serine proteinases.We reason that it is the substrate binding geometry, not the catalytic mechanism of a proteinase, that dictates its reactivity with protein inhibitors.Interleukin-1P, a potent pro-inflammatory cytokine, is synthesized as an inactive precursor that is converted to the active form by a specific proteinase, interleukin-lp converting enzyme (ICE)' (2,3).ICE is a cysteine proteinase of a previously unrecognized family, with an unusual specificity for Asp at PI (4, 51.' Based on primary sequence identity, two new members of the ICE family are now recognized, namely Ced-3 (6) and nedd-2 (7).It is not yet known whether Ced-3 or nedd-2 possess proteolytic activity, and not much is known of nedd-2 other than its transcript is down-regulated in developing mouse