Analysis of human IL-2/IL-2 receptor beta chain interactions: monoclonal antibody H2-8 and new IL-2 mutants define the critical role of alpha helix-A of IL-2.
Interleukin 2 (IL-2) interacts with a receptor (IL-2R) composed of three subunits (IL-2R alpha, IL-2R beta and IL-2R gamma). IL-2R beta plays a critical role in signal transduction. An anti-human IL-2 mAb (H2-8) produced after immunization with peptide 1-30 of IL-2 was found to recognize the region occupied by Asp20, at the exposed interface between alpha-helices A and C. Muteins at position 17 and 20 are not recognized by mAb H2-8. mAb H2-8 specifically inhibits the IL-2 proliferation of TS1beta cells which are dependent on the expression of human IL-2R beta chain for IL-2 proliferation. Substitution at internal position Leu17 demonstrates that this position is essential for IL-2 binding and IL-2 bioactivity. New IL-2 mutants at position Asp20 have been analysed. Substitutions Asp --> Asn, Asp --> Lys, Asp --> Leu, show a correlation between diminished affinity for IL-2 receptor and reduced bioactivity measured on TS1beta cells. Mutein Asp Arg lose affinity for IL-2R and bioactivity simultaneously. Furthermore, during the course of the study we have found that mutein Asp20 --> Leu is an IL-2 antagonist. The biological effects of mAb H2-8 and the properties of new mutants at positions 17 and 20 demonstrate that this region of alpha helix-A is involved in IL-2-IL-2R beta interactions.
Eckenberg
R
R
Immunogénétique Cellulaire, Institut Pasteur, Paris, France.
Xu
D
D
Moreau
J L
JL
Bossus
M
M
Mazie
J C
JC
Tartar
A
A
Liu
X Y
XY
Alzari
P M
PM
Bertoglio
J
J
Theze
J
J
D000818:Animals; D000911:Antibodies, Monoclonal; D001224:Aspartic Acid / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA; D005260:Female; D006801:Humans; D007376:Interleukin-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007930:Leucine / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding; D017433:Protein Structure, Secondary; D015375:Receptors, Interleukin-2 / Q000378:metabolism*; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D001224:Aspartic Acid; D004247:DNA; D007930:Leucine