Discrimination among rhinovirus serotypes for a variant ICAM-1 receptor molecule

Abstract

Intercellular adhesion molecule 1 (ICAM-1) is the cellular receptor for the major group of human rhinovirus serotypes, including human rhinovirus 14 (HRV14) and HRV16. A naturally occurring variant of ICAM-1, ICAM-1Kilifi, has altered binding characteristics with respect to different HRV serotypes. HRV14 binds to ICAM-1 only transiently at physiological temperatures but forms a stable complex with ICAM-1Kilifi. Conversely, HRV16 forms a stable complex with ICAM-1 but does not bind to ICAM-1Kilifi. The three-dimensional structures of HRV14 and HRV16, complexed with ICAM-1, and the structure of HRV14, complexed with ICAM-1Kilifi, have been determined by cryoelectron microscopy (cryoEM) image reconstruction to a resolution of approximately 10 angstroms. Structures determined by X-ray crystallography of both viruses and of ICAM-1 were fitted into the cryoEM density maps. The interfaces between the viruses and receptors contain extensive ionic networks. However, the interactions between the viruses and ICAM-1Kilifi contain one less salt bridge than between the viruses and ICAM-1. As HRV16 has fewer overall interactions with ICAM-1 than HRV14, the absence of this charge interaction has a greater impact on the binding of ICAM-1Kilifi to HRV16 than to HRV14.

FIG. 1.

(A) Stereo view of the surface-shaded cryoEM map of HRV16 complexed with ICAM-1 (yellow) and showing VP1 (blue), VP2 (green), and VP3 (red). The black triangle shows the limit of one icosahedral asymmetric unit. (B) Stereo view showing the quality of fit of the HRV16 capsid structure into the cryoEM map surface. VP1 (blue), VP2 (green), and VP3 (red) are represented by a trace of their C_α atoms. The density corresponding to ICAM-1 has been removed. (C) Stereo view of one ICAM-1 molecule bound to HRV16. Oneicosahedral asymmetric unit is outlined in black, showing one copy of the difference density of the ICAM-1 molecule in transparent cyan. ICAM-1 is shown as a yellow ribbon. VP1 (blue), VP2 (green), and VP3 (red) are also shown. The four Asn residues that are N-linked to carbohydrates are drawn and labeled in blue. The densities that have not been fitted with the atomic models for the ICAM-1 domains D1 and D2 are shown in red, which include the densities of the carbohydrates moieties belonging to domain D2, as well as the density of domain D3. (D) The central slab (−10 to 10 Å in the Z axis) of the cryoEM density (cyan) fitted with the appropriate backbone structures of ICAM-1 (yellow), VP1 (blue), VP2 (green), VP3 (red), and VP4 (black).

FIG. 2.

(A) Virus-receptor binding by capture ELISA. Virus was added to wells containing immobilized soluble receptor molecules or BSA (Mock) and detected by using specific primary and HRP-conjugated secondary antibodies. Bound material was visualized by the activity of HRP on the substrate 1,2-ortho-phenylene-diamine and quantified by absorbance at 490 nm. (B) Virus neutralization assays. The effect on the titer of HRV14 or HRV16 is shown after treatment with soluble receptor; antisera against HRV1B (anti-1B), HRV14 (anti-14), or HRV16 (anti-16), or BSA.

FIG. 3.

Road maps of the ICAM-1 contact area with HRV14 (left page) and HRV16 (right page). Corresponding panels show the same information for the indicated virus. Panel a is a stereo diagram of the viral surface simulated from the crystal structure coordinates within the icosahedral asymmetric unit. The rectangle indicates the area that is enlarged in panel b. The orange region in panel A corresponds to the surface region that makes contact with ICAM-1. On the left side of panels b and c are shown the surface areas of the virus or ICAM-1, respectively, with contours corresponding to separation distances of 2.0 (red), 3.0 (orange), and 4.0 (yellow) Å, representing the closest approach atoms in the receptor and in the virus. On the right side of panels b and c are shown the amino acids involved in the virus-receptor contacts. The residues are coded by their properties; positively charged (blue), negatively charged (red), polar (yellow), and hydrophobic (green) residues are shown. Shown also are the ionic networks A, B, C, and D circled by cyan dashed lines. Whereas panel b shows the surface of the virus, panel c shows the complementary surface of ICAM-1. Note the charge complementarity of the two opposing surfaces.

FIG. 3.

Road maps of the ICAM-1 contact area with HRV14 (left page) and HRV16 (right page). Corresponding panels show the same information for the indicated virus. Panel a is a stereo diagram of the viral surface simulated from the crystal structure coordinates within the icosahedral asymmetric unit. The rectangle indicates the area that is enlarged in panel b. The orange region in panel A corresponds to the surface region that makes contact with ICAM-1. On the left side of panels b and c are shown the surface areas of the virus or ICAM-1, respectively, with contours corresponding to separation distances of 2.0 (red), 3.0 (orange), and 4.0 (yellow) Å, representing the closest approach atoms in the receptor and in the virus. On the right side of panels b and c are shown the amino acids involved in the virus-receptor contacts. The residues are coded by their properties; positively charged (blue), negatively charged (red), polar (yellow), and hydrophobic (green) residues are shown. Shown also are the ionic networks A, B, C, and D circled by cyan dashed lines. Whereas panel b shows the surface of the virus, panel c shows the complementary surface of ICAM-1. Note the charge complementarity of the two opposing surfaces.

FIG. 4.

Stereo diagrams showing details of each of the ionic networks (A, B, C, and D) in the virus-receptor interface. Basic (blue), acidic (red), polar (green), and hydrophobic (gray) residues are shown. To differentiate residues in the virus to those in ICAM-1, ICAM-1 residues are enclosed in a transparent yellow van der Waals surface.

FIG. 5.

Alignment of HRV3, HRV14, HRV15, and HRV16 sequences in the vicinity of the virus-receptor contact areas. The amino acid sequence of HRV14 is closely similar to that of HRV3, as is the sequence of HRV16 to HRV15. The secondary structural elements are shown above the sequences. The “puff” region is a flexible external loop in VP2 (44). Residues in the virus-receptor interface are highlighted with colors based on the residue property. Basic (blue), acidic (red), polar (green), and hydrophobic (yellow) residues are shown. Residues that are involved in forming ionic networks are indicated by the cyan letters of A, B, C, and D, corresponding to the notation used in Fig. 3 and Table 6.