For energy supply to biomimetic constructs, a complex chemical energy-driven ATP-generating artificial system was built. The system was assembled with bottom-up detergent-mediated reconstitution of an ATP synthase and a terminal oxidase into two types of novel nanocontainers, built from either graft copolymer membranes or from hybrid graft copolymer/lipid membranes. The versatility and biocompatibility of the proposed nanocontainers was demonstrated through convenient system assembly and through high retained activity of both membrane-embedded enzymes. In the future, the nanocontainers might be used as a platform for the functional reconstitution of other complex membrane proteins and could considerably expedite the design of nanoreactors, biosensors, and artificial organelles.

cell's total energy budget. [2] The universal energy currency that is used for these purposes and can be found in all forms of life is adenosine triphosphate (ATP). A vast majority of the cellular energy demand is covered by either converting a large variety of energy-rich substrates into ATP in a process called oxidative phosphorylation, or by converting light (electromagnetic) energy into ATP in a process called photophosphorylation (or photosynthesis).These naturally existing energy conversions are valid in in the context of bottomup synthetic biology as well. In the latter, an artificial cell is envisioned as a compendium of functional modules, each hand-tailored to partially or entirely mimic one of the essential life processes, such as reproduction, growth, motility, etc. [3] Like their natural counterparts, all these reenvisioned synthetic processes are energy demanding, therefore, the deliberate design of synthetic cells should involve suitable energy management strategy, by, for example, continuous regeneration of ATP. Apart from the importance in the context of artificial cells, new energy conversion strategies can be considered as a standalone feature for enzymatic and cell-free biotechnology, wherein bottom-up synthetic biology might also deliver new and sustainable solutions.The notion of synthetic in terms of engineered and/or non-natural can be even expanded to other forms of energy (like electrical energy) and non-natural building blocks. The chemically driven ATP synthesis, as in oxidative phosphorylation, represents a spontaneous process, in which the electrons of a fuel (glucose, NADH) are transferred to an electron acceptor such as oxygen with the concomitant generation of proton gradient, which is afterwards stored again as chemical energy in ATP. In the realm of synthetic biology, other options might also be feasible, for example: can we use electrical energy and directly plug it in to drive biological processes [4,5] or make use of natural electron transfer mechanisms to produce electricity? [6] The Electron Transport Chain-a Natural Toolbox of Functional Parts for the Construction of Artificial OrganellesDuring the process of oxidative phosphorylation, electrons are passed from an electron donor ("fuel") with a more negative One of the critical steps in sustaining life-mimicking processes in synthetic cells is energy, i.e., adenosine triphosphate (ATP) regeneration. Previous studies have shown that the simple addition of ATP or ATP regeneration systems, which do not regenerate ATP directly from ADP and P i , have no or only limited success due to accumulation of ATP hydrolysis products. In general, ATP regeneration can be achieved by converting light or chemical energy into ATP, which may also involve redox transformations of cofactors. The present contribution provides an overview of the existing ATP regeneration strategies and the related nicotinamide adenine dinucleotide (NAD + ) redox cycling, with a focus on compartmentalized systems. Special attention is being paid to those approach...

Self-sustained metabolic pathways in microcompartments are the corner-stone for living systems. From a technological viewpoint, such pathways are a mandatory prerequisite for the reliable design of artificial cells functioning out-of-equilibrium. Here we develop a microfluidic platform for the miniaturization and analysis of metabolic pathways in man-made microcompartments formed of water-in-oil droplets. In a modular approach, we integrate in the microcompartments a nicotinamide adenine dinucleotide (NAD)-dependent enzymatic reaction and a NAD-regeneration module as a minimal metabolism. We show that the microcompartments sustain a metabolically active state until the substrate is fully consumed. Reversibly, the external addition of the substrate reboots the metabolic activity of the microcompartments back to an active state. We therefore control the metabolic state of thousands of independent monodisperse microcompartments, a step of relevance for the construction of large populations of metabolically active artificial cells.

Light‐driven ATP regeneration systems combining ATP synthase and bacteriorhodopsin have been proposed as an energy supply in the field of synthetic biology. Energy is required to power biochemical reactions within artificially created reaction compartments like protocells, which are typically based on either lipid or polymer membranes. The insertion of membrane proteins into different hybrid membranes is delicate, and studies comparing these systems with liposomes are needed. Here we present a detailed study of membrane protein functionality in different hybrid compartments made of graft polymer PDMS‐g‐PEO and diblock copolymer PBd‐PEO. Activity of more than 90 % in lipid/polymer‐based hybrid vesicles could prove an excellent biocompatibility. A significant enhancement of long‐term stability (80 % remaining activity after 42 days) could be demonstrated in polymer/polymer‐based hybrids.

ResumoAs obstruções das vias aéreas superiores, como tonsilas hipertrofiadas, são causas da respiração oral. Objetivo: traçar o perfil miofuncional orofacial de crianças respiradoras orais pré-adenoidectomia e/ou amidalectomia. Trata-se de um estudo descritivo de caráter quantitativo de levantamento de dados. As avaliações de motricidade orofacial basearam-se no protocolo MBGR. Resultados: foram avaliadas 32 crianças, com média de idade de 8,9 anos. Destas, 34,4% eram do sexo feminino e 65,6%, do sexo masculino; prevaleceram o aleitamento natural e o hábito oral. As principais queixas foram de respiração oral e ronco. A maioria foi encaminhada para a realização de adenoamigdalectomia. A postura corporal

A light‐driven ATP regeneration module has been created through bottom‐up assembly of small building blocks such as proteins, lipids and polymers. The use of polymer hybrid compartments instead of conventional liposomes significantly increased the long‐term stability of the module. This could play an important role especially when using the module as an energy supply to drive cell‐like functions within artificially created reaction compartments like protocells. More information can be found in the full paper by Tanja Vidaković‐Koch et al.

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