Fig. 5: Structural delineation of RUPA-58 epitope on D1.
a, Interactions between RUPA-58 and the surface of D1, as delineated by the locally refined cryo-EM map of the Pfs48/45-scTB31F:RUPA-44:RUPA-58:RUPA-154 complex. The surface representation of D1 was generated from the atomic model. b, Details of interactions between RUPA-58 and D1. D1, RUPA-58 HC, and RUPA-58 KC residues are shown in light blue, gray, and white, respectively. HCDR1, HCDR2, and HCDR3 residues are shown in pink, magenta, and purple, and KCDR1, KCDR2, and KCDR3 residues are shown in light green, green, and olive, respectively. Fab residues are annotated with H or K letters to indicate heavy and kappa light chain, respectively. The surface representation of D1 was generated from the atomic model. c, Binding kinetics of D1-directed Fabs determined using BLI with immobilized Pf D1D2 antigen. Kd is indicated along the diagonal lines. d, A heatmap of results of binding competition experiments. High signal responses for the second binding event (white) represent low competition, whereas low signal responses (black) correspond to high competition. For c and d, blue and yellow correspond to potent (IC80 < 110 μg ml–1) and non-functional mAbs (IC80 > 110 μg ml–1), respectively22. e, Epitope mapping of RUPA-71 on Pfs48/45 D1D2 (gray, ribbons), as determined by nsEM, HDX-MS, and BLI using a D1D2 construct containing p.N131Q. Left, D1D2 with the RUPA-71 epitope derived from nsEM in coral. Middle, D1D2 with the mutated residue, N131, colored in dark violet and shown as sticks. Right, D1D2 with the peptides displaying significant changes in deuterium uptake between the RUPA-71-bound D1D2 and ligand-free D1D2 as measured by HDX-MS. Small (light red) to large (dark red) decreases in deuterium uptake are shown in red; peptides that showed an increase in deuterium uptake are colored in blue. f, BLI biosensor data depicting the binding of RUPA-71 Fab (Kd = affinity) and RUPA-71 IgG (Kd = apparent affinity) to Pvs48/45–D1D2. g, RUPA-71 (gray) bound to Pvs48/45–D1, created by aligning predicted models of RUPA-71 and Pvs48/45–D1D2 generated by ABodyBuilder and RoseTTAFold38,39 to PfsD1D2 from the nsEM model. Conservation of residues in PvsD1 are shown as a gradient with identical residues in dark magenta, highly conserved residues in orchid, semi-conserved residues in pink, and non-conserved residues in light gray.
