Upon structure comparison between 11-1 p and its antagonist 11-1 ra, single or multiple residues along the 11-1 p sequence were replaced with the corresponding amino acids present in the 11-1 ra protein, in the attempt to identify sites important for receptor binding and for biologic activity on the two molecules. Ten of fifteen mutant proteins had activity comparable to that of wild-type I L - l p i n three different biologic assays and in receptor binding, indicating that the introduced changes did not influence the functional structure of the protein. Conversely, three mutants (SMIL-9: 127/263 R/T+W/Y; SMIL-10: 125/127/ 263/265 T/R/T/Q+R/W/Y/E; SMIL-15: 222/227 I/E+S/S) showed an increased binding capacity for 11-lR,, not paralleled by increased agonista ctivity, indicating that the introduced1 1-1r a residues could be involved in the nonagonist 11-1R , binding site. On the other hand, two mutants showed diminished binding capacity with concomitant decrease in biologic activity. Both mutants (SMIL-1, five substitutions in the loop 202-214; and SMIL-3, total replacement of the loop 164-173 with the 11-lra stretch 52-55) included substitutions of residues allegedly important for agonist binding to 11-1 R,. Mutant SMIL-3 showed the most profound reduction in binding capacity for 11-1 R, (CDwl21 a) and a more than 1,000-fold reduced biologic activity both in vitro and in vivo, but it retained full capacity of binding to 11-1 R,, (CDwl21 b) and acted as a selective antagonist of 11-1 R,,. From these results the following conclusions can be drawn. 11-1 p binds to 11-1 R, and to 11-1 R,, through different sites, and the loop 164-173 appears as one of the areas involved in the selective interaction with 11-lR,. Agonist (11-lp) and nonagonist (11-1 ra) binding to 11-1 R, occur through distinct sites, with loops 164-173 and 202-214 of IL-lp identified as two of the sites selectively involved in agonist binding to the activating receptor

MAPPING OF RECEPTOR-BINDING SITES ON IL-1-BETA BY RECONSTRUCTION OF IL-1RA-LIKE DOMAINS

MACKAY, ANDREW REAY;
1995-01-01

Abstract

Upon structure comparison between 11-1 p and its antagonist 11-1 ra, single or multiple residues along the 11-1 p sequence were replaced with the corresponding amino acids present in the 11-1 ra protein, in the attempt to identify sites important for receptor binding and for biologic activity on the two molecules. Ten of fifteen mutant proteins had activity comparable to that of wild-type I L - l p i n three different biologic assays and in receptor binding, indicating that the introduced changes did not influence the functional structure of the protein. Conversely, three mutants (SMIL-9: 127/263 R/T+W/Y; SMIL-10: 125/127/ 263/265 T/R/T/Q+R/W/Y/E; SMIL-15: 222/227 I/E+S/S) showed an increased binding capacity for 11-lR,, not paralleled by increased agonista ctivity, indicating that the introduced1 1-1r a residues could be involved in the nonagonist 11-1R , binding site. On the other hand, two mutants showed diminished binding capacity with concomitant decrease in biologic activity. Both mutants (SMIL-1, five substitutions in the loop 202-214; and SMIL-3, total replacement of the loop 164-173 with the 11-lra stretch 52-55) included substitutions of residues allegedly important for agonist binding to 11-1 R,. Mutant SMIL-3 showed the most profound reduction in binding capacity for 11-1 R, (CDwl21 a) and a more than 1,000-fold reduced biologic activity both in vitro and in vivo, but it retained full capacity of binding to 11-1 R,, (CDwl21 b) and acted as a selective antagonist of 11-1 R,,. From these results the following conclusions can be drawn. 11-1 p binds to 11-1 R, and to 11-1 R,, through different sites, and the loop 164-173 appears as one of the areas involved in the selective interaction with 11-lR,. Agonist (11-lp) and nonagonist (11-1 ra) binding to 11-1 R, occur through distinct sites, with loops 164-173 and 202-214 of IL-lp identified as two of the sites selectively involved in agonist binding to the activating receptor
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11697/727
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