A monoclonal antibody specific for reovirus outer-capsid protein sigma3 inhibits sigma1-mediated hemagglutination by steric hindrance.
Reovirus virions are nonenveloped icosahedral particles consisting of two concentric protein shells, termed outer capsid and core. Outer-capsid protein sigma1 is the viral attachment protein and binds carbohydrate molecules on the surface of host cells. Monoclonal antibody (MAb) 4F2, which is specific for outer-capsid protein sigma3, blocks the binding of sigma1 protein to sialic acid and inhibits reovirus-induced hemagglutination (HA). To determine whether MAb 4F2 inhibits HA by altering sigma1-sigma3 interactions or by steric hindrance, we analyzed the effect of 4F2 immunoglobulin G (IgG) and Fab fragments (Fabs) on HA induced by reovirus strain type 3 Dearing (T3D). The concentration of 4F2 IgG sufficient to inhibit T3D-induced HA was 12.5 microg per ml, whereas that of Fabs was >200 microg per ml. Dynamic light scattering analysis showed that at the concentration of IgG sufficient to inhibit HA, virion-antibody complexes were monodispersed and not aggregated. The affinity of 4F2 Fabs for T3D virions was only threefold less than that of intact IgG, which suggests that differences in HA inhibition titer exhibited by 4F2 IgG and Fabs are not attributable to differences in the affinity of these molecules for T3D virions. We used cryoelectron microscopy and three-dimensional image analysis to visualize T3D virions alone and in complex with either IgG or Fabs of MAb 4F2. IgG and Fabs bind the same site at the distal portion of sigma3, and binding of IgG and Fabs induces identical conformational changes in outer-capsid proteins sigma3 and mu1. These results suggest that MAb 4F2 inhibits reovirus binding to sialic acid by steric hindrance and provide insight into the conformational flexibility of reovirus outer-capsid proteins.
Nason
E L
EL
Department of Biochemistry and Molecular Biology, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030, USA.
Wetzel
J D
JD
Mukherjee
S K
SK
Barton
E S
ES
Prasad
B V
BV
Dermody
T S
TS
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology; D002213:Capsid / Q000276:immunology* / Q000648:ultrastructure; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000821:virology; D006386:Hemagglutination Tests; D006389:Hemagglutinins, Viral / Q000276:immunology* / Q000648:ultrastructure; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D008968:Molecular Conformation; D009500:Neutralization Tests; D012087:Reoviridae / Q000276:immunology*; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D006389:Hemagglutinins, Viral; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G