Inhibiting protein-protein interactions: a model for antagonist design.
Protein-protein interactions (PPI) are a ubiquitous mode of transmitting signals in cells and tissues. We are testing a stepwise, generic, structure-driven approach for finding low molecular weight inhibitors of protein-protein interactions. The approach requires development of a high-affinity, single chain antibody directed specifically against the interaction surface of one of the proteins to obtain structural information on the interface. To this end, we developed a single chain antibody (sc1E3) against hIL-1beta that exhibited the equivalent affinity of the soluble IL-1 receptor type I (sIL-1R) for hIL-1beta and competitively blocked the sIL-1R from binding to the cytokine. The antibody proved to be more specific for hIL-1beta than the sIL-1R in that it failed to bind to either murine IL-1beta or human/murine IL-1alpha proteins. Additionally, failure of sc1E3 to bind to several hIL-1beta mutant proteins, altered at receptor site B, indicated that the antibody interacted preferentially with this site. This, coupled with other surface plasmon resonance and isothermal titration calorimetry measurements, shows that sc1E3 can achieve comparable affinity of binding hIL-1beta as the receptor through interactions at a smaller interface. This stable single chain antibody based heterodimer has simplified the complexity of the IL-1/IL-1R PPI system and will facilitate the design of the low molecular weight inhibitors of this interaction.
Chrunyk
B A
BA
Pfizer Central Research Division, Pfizer Inc., Groton, CT 06340, USA.
Rosner
M H
MH
Cong
Y
Y
McColl
A S
AS
Otterness
I G
IG
Daumy
G O
GO
D000818:Animals; D000906:Antibodies / Q000276:immunology* / Q000494:pharmacology; D000936:Antigen-Antibody Complex / Q000032:analysis; D001667:Binding, Competitive; D002850:Chromatography, Gel; D015195:Drug Design; D006801:Humans; D007375:Interleukin-1 / Q000235:genetics / Q000276:immunology*; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D009154:Mutation; D017472:Receptors, Interleukin-1 / Q000037:antagonists & inhibitors* / Q000276:immunology; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology; D020349:Surface Plasmon Resonance; D014461:Ultracentrifugation
D000906:Antibodies; D000936:Antigen-Antibody Complex; D007375:Interleukin-1; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins