Membrane sculpting by curved DNA origami scaffolds.

Membrane sculpting and transformation is essential for many cellular functions, thus being largely regulated by self-assembling and self-organizing protein coats. Their functionality is often encoded by particular spatial structures. Prominent examples are BAR domain proteins, the 'banana-like' shapes of which are thought to aid scaffolding and membrane tubulation. To elucidate whether 3D structure can be uncoupled from other functional features of complex scaffolding proteins, we hereby develop curved DNA origami in various shapes and stacking features, following the presumable design features of BAR proteins, and characterize their ability for membrane binding and transformation. We show that dependent on curvature, membrane affinity and surface density, DNA origami coats can indeed reproduce the activity of membrane-sculpting proteins such as BAR, suggesting exciting perspectives for using them in bottom-up approaches towards minimal biomimetic cellular machineries.

Rapid folding of DNA into nanoscale shapes at constant temperature.

We demonstrate that, at constant temperature, hundreds of DNA strands can cooperatively fold a long template DNA strand within minutes into complex nanoscale objects. Folding occurred out of equilibrium along nucleation-driven pathways at temperatures that could be influenced by the choice of sequences, strand lengths, and chain topology. Unfolding occurred in apparent equilibrium at higher temperatures than those for folding. Folding at optimized constant temperatures enabled the rapid production of three-dimensional DNA objects with yields that approached 100%. The results point to similarities with protein folding in spite of chemical and structural differences. The possibility for rapid and high-yield assembly will enable DNA nanotechnology for practical applications.

Subunit Interactions and cooperativity in the microtubule-severing AAA ATPase spastin.

Spastin is a hexameric ring AAA ATPase that severs microtubules. To see whether the ring complex funnels the energy of multiple ATP hydrolysis events to the site of mechanical action, we investigate here the cooperativity of spastin. Several lines of evidence indicate that interactions among two subunits dominate the cooperative behavior: (i) the ATPase activity shows a sigmoidal dependence on the ATP concentration; (ii) ATPγS displays a mixed-inhibition behavior for normal ATP turnover; and (iii) inactive mutant subunits inhibit the activity of spastin in a hyperbolic dependence, characteristic for two interacting species. A quantitative model based on neighbor interactions fits mutant titration experiments well, suggesting that each subunit is mainly influenced by one of its neighbors. These observations are relevant for patients suffering from SPG4-type hereditary spastic paraplegia and explain why single amino acid exchanges lead to a dominant negative phenotype. In severing assays, wild type spastin is even more sensitive toward the presence of inactive mutants than in enzymatic assays, suggesting a weak coupling of ATPase and severing activity.