Evidence for conformationally different states of interleukin-10: binding of a neutralizing antibody enhances accessibility of a hidden epitope

Abstract

We present the mapping of two anti-human interleukin-10 (hIL-10) antibodies (CB/RS/2 and CB/RS/11) which have been described as binding their antigen cooperatively. The epitopes were identified using hIL-10-derived overlapping peptide scans prepared by spot synthesis. To identify residues essential for binding within the two epitopes, each position was replaced by all other L-amino acids. The epitope-derived peptides were further characterized with respect to antibody affinity and their inhibition of the antibody-hIL-10 interaction. One antibody (CB/RS/11) binds to residues which are completely buried in the X-ray structure of IL-10. Accessibility of this hidden epitope is enhanced upon binding of the antibody CB/RS/2, which recognizes a discontinuous epitope located nearby. The recognition of the hidden CB/RS/11 epitope, as well as the cooperative binding behaviour of the two antibodies, provides evidence that IL-10 can adopt a conformational state other than that observed in the crystal structure.

Is there an interaction between interleukin-10 and interleukin-22?

“…This is consistent with the presumption raised for the interaction between IL-10 and IL-10 receptor. [39][40][41] According to this, binding of IL-10 to IL-10R2 requires previous binding of IL-10 to the IL-10R1 chain, leading to the creation of a binding site for IL-10R2 through conformational change of the IL-10 molecule. Regarding IL-22, our peptide scan data suggest a similar mechanism for IL-22 as for IL-10.…”

A model of the ternary complex of interleukin-10 with its soluble receptors

“…Antibodies of group A inhibit biological activity of IL-10 in an approximately equimolar ratio, at concentrations as low as 10 pM [45]. It has been shown that monoclonal antibody CB/RS/2 recognizes two binding regions of a discontinuous epitope on the surface of the molecule that comprises the N-terminal half of helix A and helix D [46]. Thus, this antibody binds to site II of the ligand and prevents IL-10 from association with its low-affinity receptor.…”

Interleukin-10 Therapy—Review of a New Approach

“…The IL-10 receptor is composed of two different chains, ␣ (Ho et al, 1993) and ␤ (CRFB4) (Kotenko et al, 1997), both members of the class II cytokine receptor family. The interaction of hIL-10R with hIL-10 has been characterized recently and seems to be highly complex (Ho et al, 1993;Tan et al, 1993;Reineke et al, 1998Reineke et al, , 1999. The IL-10R␤ chain is essential for IL-10-mediated effects and CRFB4-deficient mice display the same phenotype as IL-10 deficient mice (Spencer et al, 1998).…”