在过去的二十年里,许多创新的进步,已经开发和优化了策略和方案以提高大肠杆菌中可溶性重组蛋白生产的质量和数量。主要挑战之一是伴侣蛋白与所需的目标重组蛋白的共洗脱。分子伴侣的去除对蛋白质产量很重要,结构测定,最佳活动,和重组蛋白的所需功能。在这一章中,我们概述了从寡聚蛋白质中去除伴侣污染物的各种策略,最终目的是改善重组蛋白的质量和正确折叠。我们已经详细讨论了全长蛋白质的纯化和表达,GNE(UDP-N-乙酰葡糖胺2-差向异构酶/N-乙酰甘露糖胺激酶),作为伴侣移除的案例研究。
In the last two decades, numerous innovative advances, strategies and protocols have been developed and optimized to improve the quality and quantity of soluble recombinant protein production in E. coli. One of the major challenges being the coelution of chaperone proteins along with desired recombinant protein of interest. The removal of
chaperones is important for protein yield, structural determination, optimal activity, and desired function of the recombinant protein. In this chapter, we outline various strategies for removal of chaperone contaminants from oligomeric proteins, with the ultimate objective of ameliorating the quality and proper folding of recombinant proteins. We have discussed in detail the purification and expression of full-length protein, GNE (UDP-N-acetylglucosamine 2-epimerase/ N-acetylmannosamine kinase), as a
case study for chaperone removal.