A PEGDA/DNA Hybrid Hydrogel for Cell-Free Protein Synthesis
A PEGDA/DNA Hybrid Hydrogel for Cell-Free Protein Synthesis
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用于无细胞蛋白质合成的 PEGDA/DNA 混合水凝胶
DOI：
10.3389/fchem.2020.00028
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发表时间：
2020-02-18
影响因子：
5.5
通讯作者：
Luo, Dan
中科院分区：
文献类型：
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作者：
Cui, Jinhui;Wu, Dan;Luo, Dan
无细胞蛋白合成（Cell-free protein synthesis， CFPS）具有快速表达蛋白质的优点，在合成生物学和蛋白质工程中得到了广泛的应用。然而，限制CFPS工业应用的关键问题是其相对较高的成本，这在一定程度上归因于一次性DNA模板的过高成本。水凝胶是一种可能的解决方案，因为它可以保存和再利用CFPS中的DNA模板，并且在提高CFPS的蛋白质产量方面具有很大的潜力。本文提出了一种低成本的聚乙二醇二丙烯酸酯（PEGDA）和DNA混合制备的水凝胶，该水凝胶能够在CFPS中高效、重复地合成蛋白质。优化了控制杂交水凝胶蛋白生成的参数。对该杂化水凝胶的结构和物理性质进行了表征。研究了溶液相体系和凝胶相体系的转录和表达动力学。结果表明，PEGDA/DNA水凝胶可以增强CFPS系统的蛋白表达，并可重复产生数十次蛋白。这种PEGDA/DNA杂交水凝胶可以作为可回收的基因载体，用于批量或连续蛋白表达，并为更强大，可扩展的蛋白生产和无细胞合成生物学铺平了道路。
Cell-free protein synthesis (CFPS) has the advantage of rapid expression of proteins and has been widely implemented in synthetic biology and protein engineering. However, the critical problem limiting CFPS industrial application is its relatively high cost, which partly attributes to the overexpense of single-use DNA templates. Hydrogels provide a possible solution because they can preserve and reutilize the DNA templates in CFPS and have great potential in elevating the protein production yield of the CFPS. Here, we presented a low-cost hybrid hydrogel simply prepared with polyethylene glycol diacrylate (PEGDA) and DNA, which is capable of high-efficient and repeated protein synthesis in CFPS. Parameters governing protein production specific to hybrid hydrogels were optimized. Structures and physical properties of the hybrid hydrogel were characterized. Transcription and expression kinetics of solution phase system and gel phased systems were investigated. The results showed that PEGDA/DNA hydrogel can enhance the protein expression of the CFPS system and enable a repeated protein production for tens of times. This PEGDA/DNA hybrid hydrogel can serve as a recyclable gene carrier for either batch or continuous protein expression, and paves a path toward more powerful, scalable protein production and cell-free synthetic biology.