Aquaporins - Expression, purification and characterization
Aquaporins - Expression, purification and characterization
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DOI：
10.1016/j.bbamem.2021.183650
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发表时间：
2021-05-23
影响因子：
3.4
通讯作者：
Hedfalk，Kristina
中科院分区：
文献类型：
--
作者：
Bill，Roslyn M.;Hedfalk，Kristina
水通道蛋白水通道促进水和小的、中性溶质在所有生命王国中沿渗透梯度的双向流动。在过去的二十年中，高质量蛋白质的可用性支持了这些水通道的结构和功能表征的进展。特别是，重组蛋白技术保证了水通道蛋白样品的供应，这些样品具有足够的质量和数量以供进一步研究。在这里，我们回顾了成功的表达，纯化和表征策略的特点，这些策略支持了这些成功，并将推动该领域的进一步突破。总的来说，大肠杆菌是原核异构体的合适宿主，而巴斯德毕赤酵母是真核变体最常用的重组宿主。一般来说，两步纯化程序是合适的后，溶解在吡喃葡萄糖苷和大多数结构是通过X射线后结晶确定。
Aquaporin water channels facilitate the bi-directional flow of water and small, neutral solutes down an osmotic gradient in all kingdoms of life. Over the last two decades, the availability of high-quality protein has underpinned progress in the structural and functional characterization of these water channels. In particular, recombinant protein technology has guaranteed the supply of aquaporin samples that were of sufficient quality and quantity for further study. Here we review the features of successful expression, purification and characterization strategies that have underpinned these successes and that will drive further breakthroughs in the field. Overall,Escherichia coliis a suitable host for prokaryotic isoforms, whilePichia pastorisis the most commonly-used recombinant host for eukaryotic variants. Generally, a two-step purification procedure is suitable after solubilization in glucopyranosides and most structures are determined by X-ray following crystallization.