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

Abstract

Background: Interleukin-10 (IL-10) is a cytokine whose main biological function is to suppress the immune response by induction of a signal(s) leading to inhibition of synthesis of a number of cytokines and their cellular receptors. Signal transduction is initiated upon formation of a ternary complex of IL-10 with two of its receptor chains, IL-10R1 and IL-10R2, expressed on the cell membrane. The affinity of IL-10R1 toward IL-10 is very high, which allowed determination of the crystal structure of IL-10 complexed with the extracellular/soluble domain of IL-10R1, while the affinity of IL-10R2 toward either IL-10 or IL-10/sIL-10R1 complex is quite low. This so far has prevented any attempts to obtain structural information about the ternary complex of IL-10 with its receptor chains.

Results: Structures of the second soluble receptor chain of interleukin-10 (sIL-10R2) and the ternary complex of IL-10/sIL-10R1/sIL-10R2 have been generated by homology modeling, which allowed us to identify residues involved in ligand-receptor and receptor-receptor interactions.

Conclusion: The previously experimentally determined structure of the intermediate/binary complex IL-10/sIL-10R1 is the same in the ternary complex. There are two binding sites for the second receptor chain on the surface of the IL-10/sIL-10R1 complex, involving both IL-10 and sIL-10R1. Most of the interactions are hydrophilic in nature, although each interface includes two internal hydrophobic clusters. The distance between C-termini of the receptor chains is 25 A, which is common for known structures of ternary complexes of other cytokines. The structure is likely to represent the biologically active signaling complex of IL-10 with its receptor on the surface of the cell membrane.

Figure 1

Stereo diagram of the superposition of sIL-10R1 and sIL-10R2. sIL-10R1 is green, sIL-10R2 is magenta, the disulfide bonds are shown in yellow.

Figure 2

Topology diagram of a single ligand domain associated with high- and low-affinity receptors. β-strands of the receptor molecules are shown as arrows and the helices are shown as rectangles. Binding loops of the receptor D1 domain are red and the loops of the D2 domain are blue. α-helices of single ligand domain are shown as circles. Helices A, B, C, and D (red) belong to one polypeptide chain of the ligand and helices E' and F' (green) belong to another polypeptide chain of the ligand. Highlighted helices (A, C, D, F') constitute the four-helix bundle that is involved in receptor binding.

Figure 3

Structure-based sequence alignment of soluble receptors sIL-10R1 and sIL-10R2. Aligned molecules have 20.4% of identical residues and 53.1% of homologous residues (red). Secondary structure elements are shown in green. Cysteines involved in disulfide bonds are highlighted in yellow, disulfide bonds are shown by black lines.

Figure 4

Stereo diagram of intermolecular hydrophobic clusters of the ternary IL-10/sIL-10R1/sIL-10R2 complex. Polypeptide chains of IL-10 are shown in orange and cyan, sIL-10R1 is green, and sIL-10R2 is magenta. Side chains of the residues that constitute hydrophobic clusters are shown in blue. Clusters 1a and 2a are the "top" and the "bottom" hydrophobic regions of IL-10/sIL-10R1 interface, clusters 1b and 2b are the "top" and the "bottom" hydrophobic regions of IL-10/sIL-10R2 interface and clusters 1c and 2c are the "top" and the "bottom" hydrophobic regions of sIL-10R1/sIL-10R2 interface. Disulfide bonds are shown in yellow. The potential glycosylation sites are shown in red.

Figure 5

Stereo diagram of the ternary complex of human IL-10/sIL-10R1/sIL-10R2. Polypeptide chains of IL-10 are shown in orange and cyan, sIL-10R1 is green, sIL-10R2 is magenta. The hypothetical cell membrane is perpendicular to the plane of the figure on the right and the twofold symmetry axis (not shown) is horizontal.

Figure 6

Stereo diagram of each interface within a single IL-10/sIL-10R1/sIL-10R2 signaling unit (panel A). Polypeptide chains of IL-10 are shown in orange and cyan. sIL-10R1 is green and sIL-10R2 is magenta. Panels B, C and D represent close-up view of each interface, including contact residues. (B)- an interface between IL-10 and sIL-10R1, (C)- an interface between IL-10 and sIL-10R2, (D)- an interface between sIL-10R1 and sIL-10R2. Intermolecular hydrogen bonds calculated with in 3.2 Å distance cutoff are shown as blue dotted lines.