The soluble expression of recombinant proteins in Escherichia coli is vital for protein applications in biotechnology and pharmaceuticals. However, the use of E. coli for efficient heterologous protein expression is hampered by several factors, such as poor expression and protein aggregation. Changing the culture or purification conditions may alleviate these issues, but methods based on gene fusion technology offer unique opportunities to improve the production and purification of soluble proteins. Here, we develop a novel fusion tag based on Spy, a newly identified molecular chaperone that functions in the periplasm of E. coli in an ATP-independent manner to prevent protein aggregation and assist in protein folding. We found that the tandem fusion of Spy stands among the well-described best fusion partners, such as MBP and SUMO, in increasing the soluble steady-state levels of six heterologous passenger proteins. Moreover, an easily aggregated passenger protein remained soluble after the removal of the Spy tag, implying that chaperone-dependent folding occurred when the passenger protein was fused to Spy. Our work expands the toolkit of fusion tags and allows them to aid in the production of unstable proteins with industrial or clinical values.

中文翻译：

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分子伴侣间谍转变为新型融合标签，以增强重组蛋白表达。

重组蛋白在大肠杆菌中的可溶性表达对于生物技术和药物中的蛋白质应用至关重要。但是，使用大肠杆菌进行有效的异源蛋白质表达受到多种因素的阻碍，例如不良的表达和蛋白质聚集。改变培养或纯化条件可以缓解这些问题，但是基于基因融合技术的方法为改善可溶性蛋白的生产和纯化提供了独特的机会。在这里，我们开发了一种基于Spy的新型融合标签，Spy是一种新鉴定的分子伴侣，它以与ATP无关的方式在大肠杆菌的周质中发挥功能，以防止蛋白质聚集并协助蛋白质折叠。我们发现Spy的串联融合是众所周知的最佳融合伙伴，例如MBP和SUMO，增加六种异源过客蛋白的可溶性稳态水平。此外，去除Spy标签后，易于聚集的客运蛋白仍保持可溶状态，这意味着将客运蛋白融合到Spy时会发生伴侣依赖性折叠。我们的工作扩展了融合标签的工具包，并使其能够协助生产具有工业或临床价值的不稳定蛋白质。