基于液滴微流控技术的蛋白质结晶

doi:10.16085/j.issn.1000-6613.2022-1074

化工进展 ›› 2023, Vol. 42 ›› Issue (4): 2024-2030.DOI: 10.16085/j.issn.1000-6613.2022-1074

基于液滴微流控技术的蛋白质结晶

吴霞(), 蒋勋涛, 张余晓, 吕慧园, 黄方, 于筱溪()

中国石油大学（华东）化学化工学院，山东青岛 266580

收稿日期:2022-06-08 修回日期:2022-08-23 出版日期:2023-04-25 发布日期:2023-05-08
通讯作者: 于筱溪
作者简介:吴霞（1997—），女，硕士研究生，研究方向为蛋白质结晶。E-mail：393602166@qq.com。
基金资助:
国家自然科学基金(22178387)

Protein crystallization research based on droplet microfluidics

WU Xia(), JIANG Xuntao, ZHANG Yuxiao, LYU Huiyuan, HUANG Fang, YU Xiaoxi()

College of Chemistry and Chemical Engineering, China University of Petroleum, Qingdao 266580, Shandong, China

Received:2022-06-08 Revised:2022-08-23 Online:2023-04-25 Published:2023-05-08
Contact: YU Xiaoxi

摘要/Abstract

摘要：

结晶是培养蛋白质单晶以及蛋白质分离纯化的一种重要手段。传统静态蛋白质结晶操作周期长，晶体粒径分布不均。液滴微流控技术能够实现低雷诺数条件下的高效传质传热，是提高蛋白质结晶速率以及改善晶体尺寸分布的潜在方法。本研究利用液滴微流控系统分别在静止液滴和低剪切速率（流动液滴）条件下进行蛋白质结晶。实验结果表明，微流控液滴内循环可以有效提高成核速率和晶体数量，并且成核速率、平均晶体数量和晶体尺寸均与液滴流速呈单调依赖关系,证明了微流控液滴内温和的剪切力场是改善蛋白质晶粒度分布，减少蛋白质晶体聚集的有效手段。此外，活性实验表明微液滴内循环对溶菌酶的活性并未产生明显影响。

关键词: 液滴微流控, 液滴内循环, 蛋白质结晶, 粒度分布

Abstract:

Crystallization is an important tool for the cultivation of protein single crystals as well as protein separation and purification. Conventional static protein crystallization has problems such as long operation cycle and uneven crystal size distribution. Droplet microfluidics is a potential method to increase protein crystallization rate and improve crystal size distribution because of its low reagent consumption and high efficiency of mass and heat transfer under low Reynolds number conditions. This study investigated lysozyme crystallization using a droplet microfluidic system under static droplet and low shear rate (flowing droplet) conditions, respectively. The experimental results showed that the microfluidic droplet internal circulation could effectively improve the nucleation rate and crystal number. Furthermore, nucleation rate, average crystal number and crystal size were monotonically dependent on the oscillation flow rate. In addition, the activity experiments showed that the forced circulation of microdroplets did not have a significant effect on the lysozyme activity.

Key words: droplet microfluidics, droplet internal circulation, protein crystallization, size distribution

中图分类号:

吴霞, 蒋勋涛, 张余晓, 吕慧园, 黄方, 于筱溪. 基于液滴微流控技术的蛋白质结晶[J]. 化工进展, 2023, 42(4): 2024-2030.

WU Xia, JIANG Xuntao, ZHANG Yuxiao, LYU Huiyuan, HUANG Fang, YU Xiaoxi. Protein crystallization research based on droplet microfluidics[J]. Chemical Industry and Engineering Progress, 2023, 42(4): 2024-2030.

图/表 13

图 1 微流控装置示意图

图 2 PMMA通道结构示意图（单位：mm）

图 3 液滴生成条件筛选结果

图4 光学显微镜实时观测液滴内晶体数量变化（45µL/min振荡流量下拍摄）

图 5 不同条件下成核速率对比图

图 6 不同条件下晶体放大图（比例尺100µm）

图 7 不同条件下晶体光学显微镜图像

图8 不同条件下晶体数量分布对比

图 9 不同条件下晶体数量对比

图 10 不同振荡条件对液滴中形成的晶体数量的影响

图 11 不同振荡条件对液滴中形成的晶体粒径的影响

图12 微液滴内流体循环示意图[20]

图13 不同流速下溶菌酶活性

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基于液滴微流控技术的蛋白质结晶

Protein crystallization research based on droplet microfluidics

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