Structure of UL18, a peptide-binding viral MHC mimic, bound to a host inhibitory receptor

Abstract

UL18 is a human cytomegalovirus class I MHC (MHCI) homolog that binds the host inhibitory receptor LIR-1 and the only known viral MHC homolog that presents peptides. The 2.2-A structure of a LIR-1/UL18/peptide complex reveals increased contacts and optimal surface complementarity in the LIR-1/UL18 interface compared with LIR/MHCI interfaces, resulting in a >1,000-fold higher affinity. Despite sharing only approximately 25% sequence identity, UL18's structure and peptide binding are surprisingly similar to host MHCI. The crystal structure suggests that most of the UL18 surface, except where LIR-1 and the host-derived light chain bind, is covered by carbohydrates attached to 13 potential N-glycosylation sites, thereby preventing access to bound peptide and association with most MHCI-binding proteins. The LIR-1/UL18 structure demonstrates how a viral protein evolves from its host ancestor to impede unwanted interactions while preserving and improving its receptor-binding site.

Fig. 1.

Structure of UL18/LIR-1 and comparison with HLA-A2/LIR-1. (A and B) Ribbon diagrams of the UL18/LIR-1 (A) and HLA-A2/LIR-1 (B) complexes. Disulfide bonds are thick yellow sticks, and ordered carbohydrates on UL18 are thin yellow sticks. A predicted O-glycosylation site in the UL18 ectodomain is indicated by an orange sphere, and the single N-glycosylation site on HLA-A2 is indicated as a yellow sphere. (C) Cα superposition of the peptide-binding platforms of UL18 and HLA-A2. Insertions in UL18 are in blue on the UL18 structure, and deletions in UL18 are black on the HLA-A2 structure. (D) The ALPHAILRL peptide model superimposed on a 2.2-Å 2F_o − F_c annealed omit electron density map contoured at 1.0 σ. (E) Stereo superposition of Cα traces of LIR-1 D1-D2 structures after superposition of the D1 domains.

Fig. 2.

Structure-based sequence alignment of UL18 and HLA-A2. Secondary structures are indicated as arrows for β-strands and springs for α-helices. Conserved residues are shown as white letters in red boxes, and conservative substitutions are indicated as red letters. UL18 residues that vary between clinical and laboratory isolates of HCMV (identified by using 21 unique HCMV sequences available in the NCBI protein database, www.ncbi.nlm.nih.gov/) are indicated with a purple dot above the UL18 sequence. Highly variable residues, defined as described previously (46), are indicated by purple dots below the HLA-A2 sequence. The asparagines in potential N-glycosylation sites in both sequences are marked with green stars. Residues in each peptide binding groove are indicated as upward-pointing black triangles. Residues that interact with LIR-1 are marked with downward-pointing blue triangles.

Fig. 3.

Sequence comparisons of residues at LIR-contacting positions. Red, blue, and yellow indicate nonconservatively substituted, conservatively substituted, and conserved positions, respectively. (A) Comparison of the LIR-1-binding (LIR-2-binding for HLA-G) epitope on the α3 domains of UL18, HLA-A2, and HLA-G. Residues contacting LIR-1 or LIR-2 (≤ 4.0 Å) are circled. Corresponding residues on class I MHC homologs that do not bind LIR-1 or LIR-2 are also listed. (B) Comparison of α3- and β2m-contacting residues on LIR-1 (LIR-2 for HLA-G) in the LIR-1/UL18, LIR-1/HLA-A2, and LIR-2/HLA-G complexes. Interacting (≤ 4.0 Å) residues are circled.

Fig. 4.

Surface representations of LIR proteins and their binding partners. Contact surfaces (≤4.0 Å) are highlighted in yellow. (A) UL18, UL18/LIR-1, and LIR-1. An asterisk marks the position of a LIR-1 loop (residues 148–154) that is disordered in the UL18/LIR-1 complex structure. (B) HLA-A2, HLA-A2/LIR-1, and LIR-1 (PDB ID code 1P7Q). (C) HLA-G, HLA-G/LIR-2, and LIR-2 (PDB ID code 2DYP).

Fig. 5.

Interaction sites for potential binding partners highlighted on a fully glycosylated UL18 model. (A) Space-filling representation of the UL18/LIR-1 complex (UL18 in magenta, β2m in slate, peptide in light green, LIR-1 in cyan) with a complex carbohydrate model (yellow) attached to each of the 13 potential N-glycosylation sites. The single predicted O-glycosylation site (residue 281) within the UL18 ectodomain is indicated by an orange sphere. Two to three additional O-glycosylation sites are predicted in the region C-terminal to the UL18 ectodomain fragment that was crystallized, but these sites would be distant from the binding sites for all potential UL18 binding partners. The UL18 counterparts of the approximate binding sites on a class I MHC molecule for a TCR or KIR are indicated by arrows. (B) Top view of the fully glycosylated UL18 peptide binding platform. (C) Potential US2 binding site (dark green) on fully glycosylated UL18. (D) Potential CD8 binding site (dark green) on fully glycosylated UL18. An enlarged view of the CD8-binding loop in the class I MHC α3 domain (gray) is shown with the counterpart UL18 loop (magenta).