Abstract Biological compartmentalization creates and controls localized environments to ensure that chemical processes are efficient, thus enabling life’s complexity and functionality. Biological systems use crystalline protein cages for nanoscale compartments, whereas larger, dynamic structures, such as vesicles and cell membranes, are formed from lipid bilayers. Although membrane-based approaches have prevailed in bottom-up synthetic biology, DNA and protein nanotechnology has focused on designing rigid cage assemblies. Here we report on the self-assembly of radially symmetric DNA origami subunits that are inspired by the structure and interactions of lipids. The formed DNA origami monolayer membranes can be readily programmed to form vesicles or hollow tubes with diameters ranging from 100 nm to over 1 μm. These DNA origami membranes represent an approach for compartmentalization that opens possibilities in bottom-up biology and cell-scale soft robotics.