Reversible membrane deformations by straight DNA origami filaments.

Membrane-active cytoskeletal elements, such as FtsZ, septin or actin, form filamentous polymers able to induce and stabilize curvature on cellular membranes. In order to emulate the characteristic dynamic self-assembly properties of cytoskeletal subunits in vitro, biomimetic synthetic scaffolds were here developed using DNA origami. In contrast to our earlier work with pre-curved scaffolds, we specifically assessed the potential of origami mimicking straight filaments, such as actin and microtubules, by origami presenting cholesteryl anchors for membrane binding and additional blunt end stacking interactions for controllable polymerization into linear filaments. By assessing the interaction of our DNA nanostructures with model membranes using fluorescence microscopy, we show that filaments can be formed, upon increasing MgCl2 in solution, for structures displaying blunt ends; and can subsequently depolymerize, by decreasing the concentration of MgCl2. Distinctive spike-like membrane protrusions were generated on giant unilamellar vesicles at high membrane-bound filament densities, and the presence of such deformations was reversible and shown to correlate with the MgCl2-triggered polymerization of DNA origami subunits into filamentous aggregates. In the end, our approach reveals the formation of membrane-bound filaments as a minimal requirement for membrane shaping by straight cytoskeletal-like objects

Shaping Giant Membrane Vesicles in 3D-Printed Protein Hydrogel Cages

Giant unilamellar phospholipid vesicles are attractive starting points for constructing minimal living cells from the bottom-up. Their membranes are compatible with many physiologically functional modules and act as selective barriers, while retaining a high morphological flexibility. However, their spherical shape renders them rather inappropriate to study phenomena that are based on distinct cell shape and polarity, such as cell division. Here, a microscale device based on 3D printed protein hydrogel is introduced to induce pH-stimulated reversible shape changes in trapped vesicles without compromising their free-standing membranes. Deformations of spheres to at least twice their aspect ratio, but also toward unusual quadratic or triangular shapes can be accomplished. Mechanical force induced by the cages to phase-separated membrane vesicles can lead to spontaneous shape deformations, from the recurrent formation of dumbbells with curved necks between domains to full budding of membrane domains as separate vesicles. Moreover, shape-tunable vesicles are particularly desirable when reconstituting geometry-sensitive protein networks, such as reaction-diffusion systems. In particular, vesicle shape changes allow to switch between different modes of self-organized protein oscillations within, and thus, to influence reaction networks directly by external mechanical cues

DNA Nanostructures on Membranes as Tools for Synthetic Biology

Over the last decade, functionally designed DNA nanostructures applied to lipid membranes prompted important achievements in the fields of biophysics and synthetic biology. Taking advantage of the universal rules for self-assembly of complementary oligonucleotides, DNA has proven to be an extremely versatile biocompatible building material on the nanoscale. The possibility to chemically integrate functional groups into oligonucleotides, most notably with lipophilic anchors, enabled a widespread usage of DNA as a viable alternative to proteins with respect to functional activity on membranes. As described throughout this review, hybrid DNA-lipid nanostructures can mediate events such as vesicle docking and fusion, or selective partitioning of molecules into phase-separated membranes. Moreover, the major benefit of DNA structural constructs, such as DNA tiles and DNA origami, is the reproducibility and simplicity of their design. DNA nanotechnology can produce functional structures with subnanometer precision and allow for a tight control over their biochemical functionality, e.g., interaction partners. DNA-based membrane nanopores and origami structures able to assemble into two-dimensional networks on top of lipid bilayers are recent examples of the manifold of complex devices that can be achieved. In this review, we will shortly present some of the potentially most relevant avenues and accomplishments of membrane-anchored DNA nanostructures for investigating, engineering, and mimicking lipid membrane-related biophysical processes

Medicina popular e feitiçaria nas visitações da Arquidiocese de Braga nos séculos XVI e XVII.

103 Jan.-Dez. 1993, p. 67-97

O Infante D. Henrique e a Colegiada da Senhora da Oliveira

A primeira visita à colegiada de GuimarãesA reacção do cabidoVisitação do Infante de 1538A terceira visitação, de 1540Análise das três visitaçõesApêndice Documenta

Membrane-coated 3D architectures for bottom-up synthetic biology dagger

One of the great challenges of bottom-up synthetic biology is to recreate the cellular geometry and surface functionality required for biological reactions. Of particular interest are lipid membrane interfaces where many protein functions take place. However, cellular 3D geometries are often complex, and custom-shaping stable lipid membranes on relevant spatial scales in the micrometer range has been hard to accomplish reproducibly. Here, we use two-photon direct laser writing to 3D print microenvironments with length scales relevant to cellular processes and reactions. We formed lipid bilayers on the surfaces of these printed structures, and we evaluated multiple combinatorial scenarios, where physiologically relevant membrane compositions were generated on several different polymer surfaces. Functional dynamic protein systems were reconstituted in vitro and their self-organization was observed in response to the 3D geometry. This method proves very useful to template biological membranes with an additional spatial dimension, and thus allows a better understanding of protein function in relation to the complex morphology of cells and organelles.We also thank the Biochemistry Core Facility of the Max Planck Institute of Biochemistry for assistance with protein purification, and the Imaging Core Facility of the same institution for assistance on the 4D image visualisation

Sociedade e delinquência no Portugal Miguelista: A arquidiocese de Braga em três devassas visitacionais de 1831

Solicitado para um artigo que se enquadrasse na homenagem ao Doutor Victor Sá na 10.ª edição da instituição do Prémio de História Contemporânea, pensei dedicar a essa publicação um artigo relativamente extenso sobre as visitações de Guilhadeses que tinha entre mãos onde estudava a Reforma católica no concelho/arciprestado de Arcos de Valdevez. Advertido posteriormente da sua reduzida extensão, resolvi então consagrar-lhe a lição de síntese das provas de agregação, proferida a 9 de Novembro de 2001 , a qual versou as relationes ad limina e os seus relatórios (1594-1640). Passando bastante tempo sem essa homenagem se concretizar em publicação, soube que a nova administração determinou limitar a temática à Época Contemporânea, devendo abordar portanto assunto posterior a 1820. Foi então que, desejando colaborar nessa homenagem, escolhi o presente tema sobre a sociedade e a delinquência arquidiocesanas no governo de D. Miguel através das devassas visitacionais

Non-Equilibrium Large-Scale Membrane Transformations Driven by MinDE Biochemical Reaction Cycles

The MinDE proteins from E. coli have received great attention as a paradigmatic biological pattern-forming system. Recently, it has surfaced that these proteins do not only generate oscillating concentration gradients driven by ATP hydrolysis, but that they can reversibly deform giant vesicles. In order to explore the potential of Min proteins to actually perform mechanical work, we introduce a new model membrane system, flat vesicle stacks on top of a supported lipid bilayer. MinDE oscillations can repeatedly deform these flat vesicles into tubules and promote progressive membrane spreading through membrane adhesion. Dependent on membrane and buffer compositions, Min oscillations further induce robust bud formation. Altogether, we demonstrate that under specific conditions, MinDE self-organization can result in work performed on biomimetic systems and achieve a straightforward mechanochemical coupling between the MinDE biochemical reaction cycle and membrane transformation.We thank MPI‐B Core Facility for assistance in protein purification