FIG. 3.
Ability of gHst to pull down gBΔ or the indicated gBH1/H8 chimeras. 293T cells were cotransfected with mixtures containing three glycoproteins, gBΔ or gBH1/H8 with gHst and gLV5 (gB+gHst+GL), or four glycoproteins, gBΔ or gBH1/H8 with gHst, gL, and gD (gB+gHst+gL+gD), as indicated at the top of the panels. The sample indicated as No gHst carried gHV5 instead of gHst (which includes the V5 epitope). Cells were harvested 16 to 20 h after transfection. Following cell lysis, gHst was allowed to react with Strep-Tactin resin. The proteins in complex with gHst were retained by the resin, separated by PAGE, and identified by Western blotting with MAb 1817 to gB, MAb V5 to gH and gL, and H170 to gD (Pull-down). A small aliquot of the lysates was separated by PAGE (Lysates), and glycoproteins were identified by WB. Plasmids encoding gHst, gHV5, and gLV5 were described previously (14). Asterisks in panels A and B identify gB4-5(H8) which was present in smaller amounts than gBΔ in the pull-down assays but not the in lysates (C and D). Squares in panels C and D indicate that gH and gL accumulated in smaller amounts in cells coexpressing gB3-4(H8). Diamonds identify small amounts of gD present in the pull-down fraction (B) in cells coexpressing gB1-3(H8) and gB3-4(H8). Triangles identify small amounts of gH/gL, in both the pull-down and lysate fractions in cells coexpressing gB5-6(H8) (E to H).