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. 2017 Dec 11;11:388. doi: 10.3389/fncel.2017.00388

Figure 2.

Figure 2

α-Synuclein dimers bind to phosphatidic acid (PA)-containing liposomes. (A) Diagram of the liposome binding assay. After incubation of α-synuclein with liposomes, ultracentrifugation in an Accudenz gradient results in flotation of liposomes along with any bound protein to top fractions. (B) Fluorometer measurements showing the distribution of liposomes after ultracentrifugation. Note that >90% of liposomes are found in fractions 1–3. (C) Western blots showing the amount of α-synuclein that is bound to liposomes (fractions 1–3) or unbound (fractions 4–8). Monomeric α-synuclein exhibits very little binding to phosphatidylcholine (PC) liposomes, but binds robustly to 1:1 PC/PA liposomes. CC and NC dimers follow a similar pattern of binding. (D) Quantification of band intensities from Western blots. Monomeric and dimeric α-synuclein all exhibit a significant increase in binding to PC/PA liposomes, compared to PC alone. Bars represent mean ± SEM of the percentage of total α-synuclein bound, as measured in n = 3–4 independent experiments. Asterisks indicate statistical significance by Student’s t-test (p < 0.05). (E) Direct comparison of PC and PC/PA binding by monomeric or dimeric α-synuclein. Asterisks indicate statistical significant by analysis of variance (ANOVA; p < 0.05).