Figure 3.

Cell-free expressed BtubA/B assemble into dynamic bMTs on SLBs. (A) Schematic of CFPS and BtubA/B polymerization on supported lipid membranes. (B) Fluorescence microscopy image of bMTs reconstituted from separately expressed BtubA and BtubB. Before the addition to an SLB the mixed solution was supplemented with a small fraction of purified Atto488-BtubA/B for imaging. Scale bar: 10 μm. (C) Cell-free synthesized BtubA was mixed with 100 nM Atto488-BtubA/B and incubated on an SLB for 30 min. No filament was observed. At time t = 0, separately expressed BtubB was added on top of the SLB, triggering the immediate formation of short filaments that developed into longer and thicker bundles. Scale bars: 10 μm.