Receptor-binding properties of a soluble form of human cytomegalovirus glycoprotein B.
The human cytomegalovirus (HCMV) glycoprotein B (gB) (also known as gpUL55) homolog is an important mediator of virus entry and cell-to-cell dissemination of infection. To examine the potential ligand-binding properties of gB, a soluble form of gB (gB-S) was radiolabeled, purified, and tested in cell-binding experiments. Binding of gB-S to human fibroblast cells was found to occur in a dose-dependent, saturable, and specific manner. Scatchard analysis demonstrated a biphasic plot with the following estimated dissociation constants (Kd): Kd1, 4.96 x 10(-6) M; Kd2, 3.07 x 10(-7) M. Cell surface heparan sulfate proteoglycans (HSPGs) were determined to serve as one class of receptors able to facilitate gB-S binding. Both HSPG-deficient Chinese hamster ovary (CHO) cells and fibroblast cells with enzymatically removed HSPGs had 40% reductions in gB-S binding, whereas removal of chondroitin sulfate had no effect. However, a significant proportion of gB-S was able to associate with the cell surface in the absence of HSPGs via an undefined nonheparin component. Binding affinity analysis of gB-S binding to wild-type CHO-K1 cells demonstrated biphasic binding kinetics (Kd1, 9.85 x 10(-6) M; Kd2, 4.03 x 10(-8) M), whereas gB-S binding to HSPG-deficient CHO-677 cells exhibited single-component binding kinetics (Kd, 7.46 x 10(-6) M). Together, these data suggest that gB-S associates with two classes of cellular receptors. The interaction of gB with its receptors is physiologically relevant, as evidenced by an inhibitory effect on HCMV entry when cells were pretreated with purified gB-S. This inhibition was determined to be manifested at the level of virus attachment. We conclude that gB is a ligand for HCMV that mediates an interaction with a cellular receptor(s) during HCMV infection.
Boyle
K A
KA
Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, 53706-1532, USA.
Compton
T
T
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003587:Cytomegalovirus / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism; D006801:Humans; D009036:Moths / Q000166:cytology; D011991:Receptors, Virus / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012995:Solubility; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*
D019812:Heparan Sulfate Proteoglycans; D011991:Receptors, Virus; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C064084:cytomegalovirus receptor; C040495:glycoprotein B, Simplexvirus