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. 2023 Apr 19;14:2219. doi: 10.1038/s41467-023-37890-2

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a Binding of CyRPA to either Rh5_unproc or Rh5_proc was determined by Biolayer Interferometry (BLI). Biotinylated CyRPA was pre-added to a Streptavidin biosensor, before dipping into twofold dilutions of Rh5_unproc and Rh5_proc at t = 0 s. Binding occurred for 120 s followed by dissociation for a further 120 s. The K_D values were calculated using Octet Data Analysis v11.0 software. b Binding of Rh5_unproc and Rh5_proc to the RCR complex by BLI. CyRPA was added at t = 0 s to biotinylated Ripr immobilised on a Streptavidin biosensor for 120 s before allowing dissociation for a further 120 s. Either Rh5_unproc or Rh5_proc were added to Ripr-CyRPA and allowed to bind, then dissociate, before PTRAMP-CSS (PC) was added at t = 480 s. Binding and dissociation were allowed to occur until t = 720 s. c Binding of Rh5_unproc or Rh5_proc and PCRC-Rh5_unproc or PCRC-Rh5_proc to erythrocytes measured by flow-cytometric analysis. The assay was done three times and means (+/−) standard error of the mean (SEM) is shown. One-way ANOVA with Sidak’s multiple comparisons test was used to calculate p values. d Representation of the binding results in c using known structures of CSS (PDB ID: 7UNZ), PfRipr, CyRPA, PfRh5 (PDB ID: 6MPV), basigin (BSG) (PDB ID: 3B5H) and the AlphaFold Monomer v2.0 structures⁵² of the PTRAMP ectodomain (Uniprot: Q8I5M8) and Rh5PD (UniProt: Q8IFM5). Structures were assembled in ChimeraX (https://www.rbvi.ucsf.edu/chimerax). Source data are provided as a source data file.