INSERTION OF A STRUCTURAL DOMAIN OF INTERLEUKIN (IL)-1-BETA CONFERS AGONIST ACTIVITY TO THE IL-1 RECEPTOR ANTAGONIST - IMPLICATIONS FOR IL-1 BIOACTIVITY

We showed previously that replacement of Lys-145 in the IL-1 receptor antagonist (IL-1ra) with Asp resulted in an analog (IL-1ra K145D) with partial agonist activity. To identify additional amino acids that aff ect IL-1 bioactivity, we created second site mutations in IL-1ra K145D , Substitutions of single amino acids surrounding position 145 were ma de; none of these substitutions increased the bioactivity of IL-1ra K1 45D. However, the insertion of the beta-bulge (QGEESN) of IL-1 beta at the corresponding region of IL-1ra K145D resulted in a 3-4-fold augme ntation of bioactivity. An additional increase in agonist activity was observed when the beta-bulge was coexpressed with a second substituti on (Kis-54 --> Pro) in IL-1ra K145D, We also show that the bioactivity of both IL-1ra K145D and the triple mutant IL-1ra K145D/H54P/QGEESN i s dependent on interaction with the newly cloned IL-1 receptor accesso ry protein.