Fig 5. LACTB filaments bind to CL-enriched liposomes.

(A–D) Representative negative-stain TEM images of LACTB filaments in the presence of liposomes. Scale bar, 100 nm. Purified LACTB filaments were incubated with CL-containing liposomes at room temperature for 4 h. Red boxes indicate protein–membrane contact sites. (E) Zoomed in views of the red boxes in (A–D), showing the details of protein–liposome interactions. (F) A schematic diagram of filament–liposome interactions illustrating different membrane-binding modes of LACTB filaments. (G) Catalytic activity of WT LACTB in the presence of liposomes. Enzyme activity was determined in vitro by using a fluorescently labeled substrate and 3 independent experiments were performed for each sample. Bars represent the mean of at least 3 independent WT and reconstitution assays. Error bars indicate standard deviation. (H) Liposome tethering activity of WT LACTB under the presence of liposomes of various compositions. Liposome tethering activity was quantified by counting 1,000 filaments from at least 3 independent reconstitution assays and calculating the total percentage of filament tethering. Bars represent the mean liposome tethering activity and error bars indicate SEM. (I) Liposome tethering activity of key residues involved in liposome binding. Liposome tethering activity was quantified by counting 1,000 filaments from at least 3 independent reconstitution assays and calculating the total percentage of filament tethering. Bars represent the mean liposome tethering activity of reconstitution assays and error bars indicate SEM. Statistical analysis was performed with an unpaired two-tailed Student t test (G) and unpaired two-tail Welch’s t tests (H, I). *p < 0.05, **p < 0.005, ns (no significance). The underlying data in (G), (H), and (I) are provided in sheets Fig 5G–5I in S1 Data. CL, cardiolipin; TEM, transmission electron microscopy; WT, wild-type.