The effects on HIV-1 infection of a glucosidase inhibitor, N-butyl deoxynojirimycin (N-buDNJ), were examined. The combinations of N-buDNJ and nucleoside analogs dideoxyinosine (DDI), dideoxycytidine (DDC), or azidothymidine (AZT) were examined in an acute infection assay. The combination of N-buDNJ and nucleoside analog reduced the yield of reverse transcriptase activity more than did either agent alone, and the effects on the number of infectious virus particles were additive or synergistic. In studies of the mechanism whereby N-buDNJ alters HIV-1 envelope fusion activity, no effects on CD4 binding were detected. However, cleavage within the V3 loop of gp120 was reduced by N-buDNJ treatment, possibly reflecting an altered conformation of this region of the envelope protein.