MODULARITY OF PROTEIN FUNCTION - CHIMERIC INTERLEUKIN-1-BETA-S CONTAINING SPECIFIC PROTEASE INHIBITOR LOOPS RETAIN FUNCTION OF BOTH MOLECULES

Although it is widely recognized that many proteins contain discrete f unctional domains, it is less certain whether smaller, less obviously discrete, units of structure will retain their specific function when transplanted into a different context. The observation that the potent inflammatory cytokine human interleukin 1beta has the same overall st ructure as soybean trypsin inhibitor (STI) (Kunitz) prompted us to rep lace a tight turn in the cytokine sequence with the large loop in soyb ean trypsin inhibitor that binds to the active site of trypsin. Wild-t ype interleukin 1beta (IL-1beta) is highly resistant to proteolysis, b ut the chimeric STI/IL is specifically cleaved by trypsin, apparently in the inserted loop. Other chimeric interleukins have also been const ructed, by replacing the same tight turn with inhibitory loops from ot her protein protease inhibitors: turkey ovomucoid inhibitor (TOI), a c hymotrypsin inhibitor, and alpha1-antitrypsin(AT), an elastase inhibit or. Although these loops come from proteins not related structurally t o interleukin 1, they confer specific protease sensitivity or inhibiti on on the chimeric cytokine. The cytokine properties of these chimeric interleukins have also been evaluated. The chimeras formed from human IL-1beta and all inhibitory loops tested bind to the interleukin 1 re ceptor with reasonable affinity. The typical cellular effects of IL-1, however, are not observed with all the recombinant proteins, thus con firming that receptor binding and signal transduction can be uncoupled . When these results are taken together with the results of site-direc ted mutagenesis of IL-1, reported in this paper and elsewhere, they al low the receptor and intracellular transduction sites on the protein t o be mapped in detail.