The high-affinity interaction between the envelope glycoprotein (gpl2O-gp4l) of the human immunodeficiency virus type 1 and its receptor, CD4, is important for viral entry into cells and therapeutical approaches based on the soluble form of CD4 (sCD4). Using flow cytometry, we studied the kinetics of binding of sCD4 to gpl20-gp41 expressed on the cell surface. sCD4 binding was dependent on sCD4 concentration and temperature and exhibited bimolecular reaction kinetics. Binding was very slow at low sCD4 concentrations (below 0.2 ,ig/ml) and low temperatures (below 13°C) but increased sharply with increasing temperature. The rate constant for association at 37°C (1.5 x 105 M-1 s-1) was 14-fold higher than at 4°C, but the affinity of sCD4 to membrane-bound gpl2O-gp4l was not significantly affected. The activation energy at higher temperatures (28 to 37°C) was less than at lower temperatures (4 to 13°C). After long periods of incubation, we observed a decrease of surface-bound sCD4 and gpl20, even at low temperatures, which was attributed to sCD4-induced shedding of gpl20. The rate of gpl20 shedding was much lower than the rate of sCD4 binding and was dependent on sCD4 concentration and temperature. The finding that sCD4 binding is slow, especially at low sCD4 concentrations, can be of critical importance for efficient blocking of viral infection by sCD4 and should be considered when designing new protocols in the therapy of AIDS patients.

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