Determination of the structure of a decay accelerating factor-binding clinical isolate of echovirus 11 allows mapping of mutants with altered receptor requirements for infection

Abstract

We have used X-ray crystallography to determine the structure of a decay accelerating factor (DAF)-binding, clinic-derived isolate of echovirus 11 (EV11-207). The structures of the capsid proteins closely resemble those of capsid proteins of other picornaviruses. The structure allows us to interpret a series of amino acid changes produced by passaging EV11-207 in different cell lines as highlighting the locations of multiple receptor-binding sites on the virion surface. We suggest that a DAF-binding site is located at the fivefold axes of the virion, while the binding site for a distinct but as yet unidentified receptor is located within the canyon surrounding the virion fivefold axes.

FIG. 1.

Scheme for generation of variant viruses. See Materials and Methods for details.

FIG. 2.

One-step growth curves of EV11-207 and variants EV11-207M, EV11-207R, and EV11-207C on Vero and HT29 cells. (A) Titers resulting from infections of Vero cells by the four viruses in the presence and absence of polyclonal anti-DAF sera. (B) Titers from infection of HT29 cells, once again in the presence and absence of polyclonal anti-DAF sera.

FIG. 3.

Infection by EV11-207 is inhibited by anti-DAF antibodies. HT29 cells were preincubated with a rabbit polyclonal sera (at a 1/500 dilution). Inhibition of CPE resulting from antibody treatment was scored 48 to 72 h after infection with EV11-207, EV11-207M, EV11-207R, or EV11-207C.

FIG. 4.

The structure of the viral protomer. (A) The structure of viral protomer is shown as a ribbon diagram to highlight secondary structural elements. Blue, VP1; green, VP2; red, VP3. Two neighboring protomers within a pentamer are shown, and the two views are related by a 90° rotation about the vertical axis (applicable also to panel B). The locations of the amino acid changes seen in variants are indicated (as shown in the key). (B) The same views as in panel A are shown with the structures of ICAM-1 and the PVR overlaid. These coordinates were obtained from the low-resolution structures of these receptors in complex with human rhinovirus 16 and human rhinovirus 14 (20) and poliovirus (3, 16). The coordinates were superposed on the EV11-207 structure by superposition of the picornavirus proteins. Both panels were drawn with Molscript (21) and rendered with Raster3D (29).

FIG. 5.

Mapping of EV11-207M, -C, and -R amino acid changes onto the surface of EV11-207. (A) Space-filling model of the complete EV11-207 virion. The majority of residues are blue, with the depth of color relating to their distance from the center of the virion. Those residues that are altered in the variants are colored as in Fig. 4. As in Fig. 4B the overlaid structures of PVR and ICAM-1 are shown to indicate the range of interactions with the viral capsid made possible by different modes of canyon-binding receptors. A single SCR domain from factor H (2) is shown in gold so that the small size of an SCR domain compared to those of the immunoglobulin (Ig) domains contained in ICAM-1 (two Ig domains contained in each coordinate set) and PVR (three Ig domains in each coordinate set) may be judged. (B and C) The canyon-binding receptors are shown more closely with views from opposite sides shown in the two panels. All panels were drawn with Molscript (21) and rendered with Raster3D (29).