Rational design of interleukin-21 antagonist through selective elimination of the gammaC binding epitope

TY - JOUR

T1 - Rational design of interleukin-21 antagonist through selective elimination of the gammaC binding epitope

AU - Kang, Lishan

AU - Bondensgaard, Kent

AU - Li, Tengkun

AU - Hartmann, Rune

AU - Hjorth, Siv A

PY - 2010/4/16

Y1 - 2010/4/16

N2 - The cytokine interleukin (IL)-21 exerts pleiotropic effects acting through innate as well as adaptive immune responses. The activities of IL-21 are mediated through binding to its cognate receptor complex composed of the IL-21 receptor private chain (IL-21Ralpha) and the common gamma-chain (gammaC), the latter being shared by IL-2, IL-4, IL-7, IL-9, and IL-15. The binding energy of the IL-21 ternary complex is predominantly provided by the high affinity interaction between IL-21 and IL-21Ralpha, whereas the interaction between IL-21 and gammaC, albeit essential for signaling, is rather weak. The design of IL-21 analogues, which have lost most or all affinity toward the signaling gammaC chain, while simultaneously maintaining a tight interaction with the private chain, would in theory represent candidates for IL-21 antagonists. We predicted the IL-21 residues, which compose the gammaC binding epitope using homology modeling and alignment with the related cytokines, IL-2 and IL-4. Next we systematically analyzed the predicted binding epitope by a mutagenesis study. Indeed two mutants, which have significantly impaired gammaC affinity with undiminished IL-21Ralpha affinity, were successfully identified. Functional studies confirmed that these two novel hIL-21 double mutants do act as hIL-21 antagonists.

AB - The cytokine interleukin (IL)-21 exerts pleiotropic effects acting through innate as well as adaptive immune responses. The activities of IL-21 are mediated through binding to its cognate receptor complex composed of the IL-21 receptor private chain (IL-21Ralpha) and the common gamma-chain (gammaC), the latter being shared by IL-2, IL-4, IL-7, IL-9, and IL-15. The binding energy of the IL-21 ternary complex is predominantly provided by the high affinity interaction between IL-21 and IL-21Ralpha, whereas the interaction between IL-21 and gammaC, albeit essential for signaling, is rather weak. The design of IL-21 analogues, which have lost most or all affinity toward the signaling gammaC chain, while simultaneously maintaining a tight interaction with the private chain, would in theory represent candidates for IL-21 antagonists. We predicted the IL-21 residues, which compose the gammaC binding epitope using homology modeling and alignment with the related cytokines, IL-2 and IL-4. Next we systematically analyzed the predicted binding epitope by a mutagenesis study. Indeed two mutants, which have significantly impaired gammaC affinity with undiminished IL-21Ralpha affinity, were successfully identified. Functional studies confirmed that these two novel hIL-21 double mutants do act as hIL-21 antagonists.

KW - Amino Acid Sequence

KW - Amino Acid Substitution

KW - Binding Sites

KW - Cell Line

KW - Drug Design

KW - Epitopes

KW - Humans

KW - Interleukin-2

KW - Interleukin-4

KW - Interleukins

KW - Models, Molecular

KW - Molecular Sequence Data

KW - Multiprotein Complexes

KW - Mutagenesis, Site-Directed

KW - Protein Interaction Domains and Motifs

KW - Receptors, Interleukin-21

KW - Recombinant Proteins

KW - Sequence Homology, Amino Acid

KW - Structural Homology, Protein

U2 - 10.1074/jbc.M110.101444

DO - 10.1074/jbc.M110.101444

M3 - Journal article

C2 - 20167599

SN - 0021-9258

VL - 285

SP - 12223

EP - 12231

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 16

ER -