The promolion of membrzme fusion by the fusion (F) protein of human parathfluenza vires 3 (fiPIV3) is dependenl on a vies specific eontnbution from the fiemagglulinin-neuraminidase (HN) protein By evaluarion of chimeric hPIV3-Newcasfle disease vires (NDV) HN proteins, we have previously show n that hPl V3 F-speci ficity is determined by a domain that extends lYnm the middle of the membrane anchor to the 82rid residue in the ectodomain [Virology 209. (1995) 457; Arch Virol 13 (1997) 115]. If the corresponding NDV-derived residues replace the two C-terminat residues in this domain, no fusion is dcteeled. However. these substitutions restore a glycosyladon sile present in NDV HN, but not in hPIV3 HN. Deletion of this site from a nested set of chbneras with fiPIV3-derived N terminal porlions of decreasing length partially restores fusion, suggesting that an ofigosacchande near the top of hPIV3 HN stalk modulates fusion In addition, further mutational analyses show that a chimera with only 125 N temdnal hPIV3-dedved residues (72 in the stalk) actually promotes fusion more efficiently than Ihe wt protein These findings Iocalize the C ternrinus of the F-specific domain in hPlV3 HN n full 10 residues closer to the membrane than previously shown