Journal: Nature Communications

Article Title: De novo synthesized Min proteins drive oscillatory liposome deformation and regulate FtsA-FtsZ cytoskeletal patterns

doi: 10.1038/s41467-019-12932-w

Figure Lengend Snippet: Supported membrane assays with de novo synthesized MinD and MinE proteins. a Schematic of the experimental workflow for end-point expression assays. Both minD and minE genes are expressed in a PURE system reaction in the presence of DnaK chaperone mix for 3 h. The solution is then supplemented with 2.5 mM ATP and a trace amount of purified eGFP-MinD (100 nM) before transfer on top of a supported lipid bilayer. b Fluorescence microscopy images of representative types of MinDE dynamic patterns. Videos can be found in Supplementary Movie . Scale bars are 20 µm. c Schematic illustration of the in situ MinDE co-expression and self-organization on an SLB. d Several SLB fields of view were imaged at different points during in situ co-expression of MinD and MinE proteins. Corresponding videos are shown in Supplementary Movie . Scale bars are 20 µm

Article Snippet: The solution was supplemented with (final concentrations indicated): 2.5 mM ATP, 100 nM purified eGFP-MinD, DnaK (GeneFrontier Corporation), and MilliQ water to adjust the final volume to 20 µL.

Techniques: Synthesized, Expressing, Purification, Fluorescence, Microscopy, In Situ

Journal: Nature Communications

Article Title: De novo synthesized Min proteins drive oscillatory liposome deformation and regulate FtsA-FtsZ cytoskeletal patterns

doi: 10.1038/s41467-019-12932-w

Figure Lengend Snippet: In-liposome production and self-organization of MinDE proteins. a Schematic illustration of liposome reconstitution assays. The minD and minE genes are co-expressed within phospholipid vesicles in the presence of DnaK chaperones, extra ATP, and 1 µM of purified eGFP-MinC to report for MinDE dynamics. b Time-series images showing liposomes (PC/PG composition) exhibiting periodic redistribution of eGFP-MinC between the membrane and the lumen (Supplementary Movie ). Colored arrowheads point to Min-active liposomes. Other liposomes show stable fluorescence signal at the membrane or in the lumen throughout the image sequence. Scale bars are 10 µm. c Zoomed-in images taken at three different time points (from left to right) of liposomes (PC/PG composition) exhibiting the pulsing, pole-to-pole, or circling modes of oscillations (Supplementary Movie ). The corresponding kymographs are displayed below. Scale bars are 10 µm. d Time traces of the fluorescence intensity of eGFP-MinC at the membrane (magenta) and in the lumen (green) for the three liposomes shown in c . a.u., arbitrary unit. e The effects of lipid composition and vesicle size have been analyzed for liposomes with a Min-pulsing behavior. Both PC/PG (black dots) and PC/PG/PE/CL (green dots) liposomes show a cross-section area (averaged over the different time point images) that is not obviously correlated with the period of oscillations. Twenty liposomes of each composition have been analyzed

Article Snippet: The solution was supplemented with (final concentrations indicated): 2.5 mM ATP, 100 nM purified eGFP-MinD, DnaK (GeneFrontier Corporation), and MilliQ water to adjust the final volume to 20 µL.

Techniques: Purification, Fluorescence, Sequencing