The 369-residue glycoprotein D (gD) is the entry, receptor-binding protein of herpes simplex virus 1. The common receptors for viral entry are nectin-1, HveA, and a specific O-linked sulfated proteoglycan. The major receptor-binding sites of gD are at the N terminus, whereas the domain required for fusion of viral envelope with the plasma membrane is at the C terminus of the ectodomain (residues 260-310). In the course of retargeting gD to the urokinase plasminogen activator (uPA) receptor for potential therapeutic applications, we obtained a genetically engineered infectious virus in which the receptor-binding domain consisting of the N-terminal domain of uPA fused to residues 33-60 of gD was separated from an independently expressed C-terminal domain of gD containing residues 219-369. The intervening sequences (residues 62-218) were replaced by a stop codon and a promoter for the C-terminal domain of gD. The physical interaction of the two components was reconstructed by coimmunoprecipitation of the N-terminal domain of uPA with the C-terminal domain of gD. These results indicate that codons 61-218 of gD do not encode executable functions required for viral entry into cells and suggest that the receptor-binding ligand must interact with but need not alter the structure of the residual portion of gD to effect virus entry. This finding opens the way for the development of a family of recombinant viruses in which the profusion domain of gD and independently furnished, interacting receptor-binding domains effect entry of the virus via a range of receptors.