Department of Cellular Biophysics, Max Planck Institute for Medical Research, Jahnstraße 29, D-69120 Heidelberg, Germany.
Institute for Molecular Systems Engineering (IMSE), Heidelberg University, Im Neuenheimer Feld 225, D-69120 Heidelberg, Germany.
ACS Synth Biol. 2022 Jan 21;11(1):366-382. doi: 10.1021/acssynbio.1c00472. Epub 2021 Dec 10.
By using electrostatic interactions as driving force to assemble vesicles, the droplet-stabilized method was recently applied to reconstitute and encapsulate proteins, or compartments, inside giant unilamellar vesicles (GUVs) to act as minimal synthetic cells. However, the droplet-stabilized approach exhibits low production efficiency associated with the troublesome release of the GUVs from the stabilized droplets, corresponding to a major hurdle for the droplet-stabilized approach. Herein, we report the use of pH as a potential trigger to self-assemble droplet-stabilized GUVs (dsGUVs) by either bulk or droplet-based microfluidics. Moreover, pH enables the generation of compartmentalized GUVs with flexibility and robustness. By co-encapsulating pH-sensitive small unilamellar vesicles (SUVs), negatively charged SUVs, and/or proteins, we show that acidification of the droplets efficiently produces dsGUVs while sequestrating the co-encapsulated material. Most importantly, the pH-mediated assembly of dsGUVs significantly improves the production efficiency of free-standing GUVs (i.e., released from the stabilizing-droplets) compared to its previous implementation.
利用静电相互作用作为驱动力来组装囊泡，最近采用液滴稳定法将蛋白质或隔室重新构建并封装在巨大的单层囊泡（GUV）中，充当最小的合成细胞。然而，液滴稳定方法的生产效率低，与从稳定液滴中释放 GUV 相关的麻烦有关，这对应于液滴稳定方法的主要障碍。在此，我们报告了使用 pH 值作为通过体相或基于液滴的微流控技术自组装液滴稳定的 GUV（dsGUV）的潜在触发因素。此外，pH 值能够生成具有灵活性和鲁棒性的隔室化 GUV。通过共包封 pH 敏感的小单层囊泡（SUV）、带负电荷的 SUV 和/或蛋白质，我们表明液滴酸化可以有效地生成 dsGUV，同时将共包封的物质隔离。最重要的是，与之前的实施方式相比，pH 介导的 dsGUV 组装显著提高了游离-standing GUV（即从稳定液滴中释放）的生产效率。
