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Comparative Study
. 2009 Jun;83(11):5556-66.
doi: 10.1128/JVI.02532-08. Epub 2009 Mar 25.

Structural comparison of different antibodies interacting with parvovirus capsids

Susan Hafenstein  1 , Valorie D Bowman, Tao Sun, Christian D S Nelson, Laura M Palermo, Paul R Chipman, Anthony J Battisti, Colin R Parrish, Michael G Rossmann
Affiliations
Comparative Study

Structural comparison of different antibodies interacting with parvovirus capsids

Susan Hafenstein et al. J Virol. 2009 Jun.

Abstract

The structures of canine parvovirus (CPV) and feline parvovirus (FPV) complexed with antibody fragments from eight different neutralizing monoclonal antibodies were determined by cryo-electron microscopy (cryoEM) reconstruction to resolutions varying from 8.5 to 18 A. The crystal structure of one of the Fab molecules and the sequence of the variable domain for each of the Fab molecules have been determined. The structures of Fab fragments not determined crystallographically were predicted by homology modeling according to the amino acid sequence. Fitting of the Fab and virus structures into the cryoEM densities identified the footprints of each antibody on the viral surface. As anticipated from earlier analyses, the Fab binding sites are directed to two epitopes, A and B. The A site is on an exposed part of the surface near an icosahedral threefold axis, whereas the B site is about equidistant from the surrounding five-, three-, and twofold axes. One antibody directed to the A site binds CPV but not FPV. Two of the antibodies directed to the B site neutralize the virus as Fab fragments. The differences in antibody properties have been linked to the amino acids within the antibody footprints, the position of the binding site relative to the icosahedral symmetry elements, and the orientation of the Fab structure relative to the surface of the virus. Most of the exposed surface area was antigenic, although each of the antibodies had a common area of overlap that coincided with the positions of the previously mapped escape mutations.

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Figures

FIG. 1.
FIG. 1.
The viral surface is shown as a stereographic projection where the polar angles φ and θ represent the latitude and longitude of a point on the viral surface, respectively (63). The virus surface is represented as a quilt of amino acids (52), and the icosahedral asymmetric unit of the virus is indicated by the triangular boundary. To visualize the different copies of the capsid protein in the road map that comprise the asymmetric unit, VP2 molecules are displayed in tints of magenta, green, and blue. Escape mutations from neutralizing antibodies are colored red for site A and dark blue for site B.
FIG. 2.
FIG. 2.
The crystal structure of Fab 14 is shown in stereo as a ribbon diagram. The CDRs are labeled for the heavy chain (cyan) and the light chain (magenta).
FIG. 3.
FIG. 3.
Surface-rendered cryoEM reconstructions of virus complexed with each of eight fragments from neutralizing antibodies described by Nelson et al. (35). Surfaces are shown at one sigma of the cryoEM density. Density further than a 125-Å radius from the center of the virus is shown in gray. Shown directly below each reconstruction is the footprint of the Fab in the complex. Both complexes and footprints are color coded red for antibodies directed to the A site and blue for antibodies directed to the B site.
FIG. 4.
FIG. 4.
Amino acid sequences of the light and heavy chains aligned according to consensus sequences (red) and CDR loops (blue). Residues in contact with the viral surface are indicated by asterisks. Since only the light chain sequence was determined for Fab 16, the mouse antibody 1AIF Fab was used to interpret the Fab 16 density.
FIG. 5.
FIG. 5.
The viral surface is shown as a stereographic projection where the polar angles φ and θ represent the latitude and longitude of a point on the viral surface, respectively (63). (A) The residues shared by the Fab footprints are defined as the common A site and common B site and outlined in red and blue, respectively. (B) All of the residues within the antibody footprints define a total antigenic surface outlined in black. The viral surface residues are colored according to the distance from the center of the virus, with red amino acids being the furthest away and blue representing depressions in the viral surface.
FIG. 6.
FIG. 6.
Capsid protein of CPV viewed approximately tangentially to the surface of the virus, showing the closest icosahedral five- and threefold symmetry axes. Spheres show the positions of residues previously identified as belonging to the A site (red) and B site (blue). Additional residues found in the footprints of all antibodies directed to the A site are shown in magenta, and residues common to all B site antibodies are shown in cyan. Secondary structural elements, or β strands, are labeled A to I.
FIG. 7.
FIG. 7.
The Fab footprints for the A site (red X's) and the B site (blue X's) overlap with the TfR binding site, identified in black by amino acid number (19). The antibodies 8, 15, and 16 directed to the B site show the greatest amount of overlap with the receptor binding site.
FIG. 8.
FIG. 8.
The cryoEM density of the CPV-Fab 14 complex shown as a surface-contoured mesh in a slab view at the region of the threefold spike. The icosahedral threefold axis is indicated by a dark gray axis. The heavy chain (green) and the light chain (gold) of Fab 14 are fitted into the Fab density. Both CPV (blue) and FPV (magenta) have been fitted into the viral density to show the very subtle differences between the two viruses at loops 1 and 2 in the interface of the capsid interactions with the Fab.
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