Fig. 2
From: Engineered botulinum neurotoxin B with improved efficacy for targeting human receptors
Combinational mutations in HCB enhance its binding to h-Syt II and Syt I. a HCB double mutations were examined in pull-down assays for their abilities to bind m-Syt II vs. h-Syt II. Three double mutations that showed robust binding to h-Syt II are marked in red. One of two independent experiments is shown. b Binding of HCB to h-Syt II was quantified by the BLI assay. Representative association and dissociation curves are shown for WT HCB and three HCB mutants: E1191M/S1199Y, E1191V/S1199W, and E1191C/S1199W. Binding parameters for all 12 double mutations are listed in Table 1, and their representative binding traces are shown in Supplementary Fig. 3. c Binding of WT HCB, HCB (E1191M), and HCB (E1191M/S1199Y) to immobilized GST-tagged h-Syt I (residues 1–80) was examined in pull-down assays, with (+) or without (−) gangliosides (Gangl.). Binding of WT HCB to h-Syt I requires the presence of gangliosides, while HCB (E1191M) and HCB (E1191M/S1199Y) bind to h-Syt I in the absence of gangliosides. One of two independent experiments is shown. Further analysis of HCB binding to h-Syt I by the BLI assay is shown in Supplementary Fig. 4 and Table 1
