FIGURE 2.

Role of the β6S4/β1H1 region of G in membrane fusion and Mab45 binding enhancement. A, schematic of NiV-G showing the various domains. Deletion sites 1, 2, 3, 4, 9, 11, and 14 are depicted, and their sequences are shown in supplemental Table 1. B, binding of anti-HA, Mab26, and Mab45 to CHO cells expressing WT or deletion mutant G constructs Δ4 and Δ9, as measured by flow cytometry. Average ± S.E. are shown, n = 3. Both Mab45 and Mab26 are conformational antibodies, as they do not Western blot. C, sequence of region 4 of G showing the location of the triple Ala mutations 4-1, 4-2, 4-3, 4-4, and 4-5 constructed in the context of full-length G. D, binding of the full-length WT and the indicated mutant G proteins to Mab26 and Mab45 by flow cytometry, normalized to their anti-HA antibody binding levels. Average ± S.E. are shown, n = 3. E, levels of binding of region 4 mutants to polyclonal anti-NiV-G antiserum 806 or to soluble ephrinB2 (B2) in CHO cells and levels of syncytia formation in 293T cells normalized to anti-HA antibody binding to correct for any discrepancies in cell surface expression levels. Average ± S.E. values are shown, n = 3. F, levels of binding of WT and region 4 mutant proteins to Mab45 at various concentrations of B2, normalized to Mab45 binding in the absence of B2 (set at 100%) as in Fig. 1C. Data were plotted using Prism, and average ± S.E. values are shown, n = 3.