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. 2019 May 30;10:2370. doi: 10.1038/s41467-019-10345-3

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© The Author(s) 2019

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. 6 — RHD structure determines in vitro membrane binding and liposome remodeling activity. a Liposome co-flotation assay to evaluate membrane-binding properties of FAM134B-RHD and various deletion mutants (see the section “Methods”). Purified protein samples were incubated with liposomes for 2 h at 37 °C and subjected to flotation on a sucrose cushion (top to bottom, 1–8) followed by SDS–PAGE and western blotting with anti-GST antibody. b–i Representative nsTEM micrographs of remodeled proteoliposomes (scale bars, 200 nm). Empty liposomes (b) were remodeled by incubation after addition of purified (c) GST, (d) wild-type RHD, (e) ΔAH_L + AH_C, (f) ΔTM12, (g) ΔTM34, (h) ΔTM12 + TM34, and (i) RHD_143–260 for 18 h at 22 °C. Insets (red squares); magnified micrographs showing examples of representative proteoliposomes with diameters measured (dotted lines) using ImageJ. j Violin plots show the measured proteoliposome size-distributions (n = 300 each) from nsTEM images. Violins shows a central boxplot (median with interquartile range, black lines) along with mirrored histograms on either sides (colored)