FIG 5.

First generation of engineered Env proteins for enhanced presentation of the PG16-recognized conformation. (A) Expi293F cells expressing QES variants containing mutations to subunit surfaces (magenta), or additional mutations to core residues (blue), bind more PG16 than wild-type Env (gray). Env variants were tested from five HIV-1 strains (from left to right: BaL, Q769.d22, Q842.d12, 25711, and DU422). The data are means ± the SD (n = 3 to 4). (B) Binding of CD4(D1-D2) to cells expressing Env variants. Inclusion of the V255M core mutation in constructs BaL-QES.i01.c01, Q769.d22-QES.i03.V255M, and DU422-QES.c03 reduces CD4 binding. The data are means (n = 2), with error bars showing the range. (C to E) Binding of PGT121 (C), PGT128 (D), and PGT145 (E) to transfected cells expressing wild-type or QES variant Env sequences. The data are means (n = 2), with error bars showing the range. (F) Wild-type and QES variants of Env were expressed with N-terminal c-myc tags. Total surface expression (gray; detected using fluorescent anti-myc staining) and PG16 binding (magenta; 2 nM PG16) to the same transfected cell samples were measured by flow cytometry. Mean fluorescence is normalized to cells expressing the respective wild-type Env. The data are means ± the SD (n = 4). Expression of BaL Env with an N-terminal c-myc tag was not detected and is not plotted.