溴化1-辛基-3-甲基咪唑聚集状态对蛋白质结晶的影响研究

doi:10.11949/0438-1157.20210214

化工学报 ›› 2021, Vol. 72 ›› Issue (9): 4854-4860.DOI: 10.11949/0438-1157.20210214

溴化1-辛基-3-甲基咪唑聚集状态对蛋白质结晶的影响研究

于筱溪¹(),闫真真¹,蒋其辉²,吴霞¹,张余晓¹,王晓娟¹,黄方^1,³

^1.中国石油大学（华东）化学工程学院，山东青岛 266580
^2.中国石油集团工程技术研究院有限公司井下作业研究所，北京 102206
^3.中国石油大学（华东）重质油国家重点实验室，山东青岛 266580

收稿日期:2021-02-04 修回日期:2021-05-07 出版日期:2021-09-05 发布日期:2021-09-05
通讯作者: 于筱溪
作者简介:于筱溪(1988—)，女，博士，副教授，yuxiaoxi@upc.edu.cn
基金资助:
中国石油集团工程技术研究院有限公司开放课题(F2020185)

Study on the effect of 1-octyl-3-methylimidazole bromide aggregation state on protein crystallization

Xiaoxi YU¹(),Zhenzhen YAN¹,Qihui JIANG²,Xia WU¹,Yuxiao ZHANG¹,Xiaojuan WANG¹,Fang HUANG^1,³

^1.College of Chemical Engineering, China University of Petroleum (East China), Qingdao 266580, Shandong, China
^2.Downhole Operation Research Department, CNPC Engineering Technology R&D Company Limited, Beijing 102206, China
^3.State Key Laboratory for Heavy Oil Processing, China University of Petroleum (East China), Qingdao 266580, Shandong, China

Received:2021-02-04 Revised:2021-05-07 Online:2021-09-05 Published:2021-09-05
Contact: Xiaoxi YU

摘要/Abstract

摘要：

研究了离子液体溴化1-辛基-3-甲基咪唑（［C₈mim］Br）添加剂的聚集状态对蛋白质结晶过程的影响。实验发现［C₈mim］Br可以改善溶菌酶晶体形貌，影响晶体数量和晶体尺寸。通过测定不同浓度［C₈mim］Br下溶菌酶的溶解度、结晶动力学、聚集状态和ζ-电势，揭示了离子液体［C₈mim］Br对溶菌酶结晶的影响机理。结果表明，［C₈mim］Br与溶菌酶的作用方式随［C₈mim］Br的聚集状态而改变。当［C₈mim］Br浓度较低（<0.1 mol/L）时，［C₈mim］⁺与溶菌酶分子之间存在疏水作用，可以促进蛋白质分子聚集，促进结晶过程。当［C₈mim］Br浓度较高（>0.1 mol/L）时，［C₈mim］⁺自发聚集成胶束，并与蛋白质分子相互作用形成溶菌酶-胶束复合物；动态光散射（DLS）结果显示结晶过程以溶菌酶-胶束复合物为基本单元聚集，因此结晶过程减缓，晶体质量提高。

关键词: 离子液体, 聚集, 蛋白质, 结晶

Abstract:

The effect of the aggregation state of the ionic liquid 1-octyl-3-methylimidazole bromide([C₈mim]Br) additives on the protein crystallization process was studied. Hen egg white lysozyme was chosen as the model protein and [C₈mim]Br was applied as the additive. It was found that the presence of [C₈mim]Br could improve the crystal morphology as well as affect the number and size of crystals. The influence mechanism of ionic liquid [C₈mim]Br on the crystallization of lysozyme was revealed by measuring the solubility, crystallization kinetics, aggregation state and zeta-potential at different [C₈mim]Br concentrations. The results showed that the interaction mechanism between lysozyme and [C₈mim]Br varied with the aggregation states of [C₈mim]Br. When the concentration of [C₈mim]Br was low (< 0.1 mol/L), there was hydrophobic interactions between [C₈mim]+ and lysozyme molecules, which promoted the aggregation and crystallization process. When the concentration of [C₈mim]Br was high (> 0.1 mol/L), [C₈mim]+ self-aggregated into micelle and formed aggregate complex with lysozyme molecules. The results from DLS further showed that the lysozyme-micelle complex was the basic unit in nucleation and hence, a slower crystallization rate and a higher crystal quality were obtained.

Key words: ionic liquids, aggregation, protein, crystallization

中图分类号:

TQ 423.9

于筱溪, 闫真真, 蒋其辉, 吴霞, 张余晓, 王晓娟, 黄方. 溴化1-辛基-3-甲基咪唑聚集状态对蛋白质结晶的影响研究[J]. 化工学报, 2021, 72(9): 4854-4860.

Xiaoxi YU, Zhenzhen YAN, Qihui JIANG, Xia WU, Yuxiao ZHANG, Xiaojuan WANG, Fang HUANG. Study on the effect of 1-octyl-3-methylimidazole bromide aggregation state on protein crystallization[J]. CIESC Journal, 2021, 72(9): 4854-4860.

图/表 8

图1 溶菌酶晶体显微镜形貌图（比例尺:100 μm）溶菌酶浓度20 mg/ml；氯化钠浓度4%；［C8mim］Br浓度/（mol/L）：（a） 0；（b） 0.02；（c） 0.06；（d） 0.1；（e） 0.3；（f） 0.7

Fig.1 Microgscope photos of lysozyme crystals (scale bars：100 μm)

图2 不同浓度[C8mim]Br下溶菌酶晶体数量

Fig.2 Numbers of lysozyme crystals under different [C8mim]Br concentration

图3 不同浓度[C8mim]Br下溶菌酶活性

Fig.3 Lysozyme activity ratio under different [C8mim]Br concentration

图4 不同浓度[C8mim]Br下溶菌酶溶解度

Fig.4 Solubility of lysozyme under different [C8mim]Br concentration

图5 不同浓度[C8mim]Br下溶菌酶结晶动力学

Fig.5 Lysozyme crystallization kinetics under different [C8mim]Br concentration

图6 不同浓度[C8mim]Br下溶菌酶聚集体ζ-电势

Fig.6 The ζ-potential of lysozyme under different [C8mim]Br concentration

图7 [C8mim]Br溶液中聚集体粒径分布(a);结晶溶液中聚集体粒径分布(b)

Fig.7 Particle size distribution of aggregates in [C8mim]Br solution (a); Particle size distribution of aggregates in crystallization solution with protein and [C8mim]Br (b)

图8 成核阶段聚集体尺寸变化溶菌酶浓度20 mg/ml；A—［C8mim］Br浓度0.3 mol/L，氯化钠浓度4%；B—氯化钠浓度4%；C—［C8mim］Br浓度0.3 mol/L；D—［C8mim］Br浓度0.1 mol/L

Fig.8 Aggregates change during induction period

参考文献 35

1	Hartje L F, Snow C D. Protein crystal based materials for nanoscale applications in medicine and biotechnology[J]. Wiley Interdisciplinary Reviews: Nanomedicine and Nanobiotechnology, 2019, 11(4): e1547.
2	Blundell T L. Protein crystallography and drug discovery: recollections of knowledge exchange between academia and industry[J]. IUCrJ, 2017, 4(pt 4): 308-321.
3	Moreno A. New trends in protein crystallization and protein crystallography[J]. Crystals, 2020, 10(1): 46.
4	Bujacz A, Talaj J A, Zielinski K, et al. Crystal structures of serum albumins from domesticated ruminants and their complexes with 3,5-diiodosalicylic acid[J]. Acta Crystallographica Section D, Structural Biology, 2017, 73(pt 11): 896-909.
5	Yamazaki T, Van Driessche A E S, Kimura Y. High mobility of lattice molecules and defects during the early stage of protein crystallization[J]. Soft Matter, 2020, 16(8): 1955-1960.
6	McPherson A, Nguyen C, Cudney R, et al. The role of small molecule additives and chemical modification in protein crystallization[J]. Crystal Growth & Design, 2011, 11(5): 1469-1474.
7	Schröder C. Proteins in ionic liquids: current status of experiments and simulations[J]. Topics in Current Chemistry, 2017, 375(2): 1-26.
8	Singh K, Marangoni D G, Quinn J G, et al. Spontaneous vesicle formation with an ionic liquid amphiphile[J]. Journal of Colloid and Interface Science, 2009, 335(1): 105-111.
9	Chen X W, Liu J W, Wang J H. Ionic liquids in the assay of proteins[J]. Analytical Methods, 2010, 2(9): 1222-1226.
10	Chen X W, Ji Y P, Wang J H. Improvement on the crystallization of lysozyme in the presence of hydrophilic ionic liquid[J]. Analyst, 2010, 135(9): 2241-2248.
11	Judge R A, Takahashi S, Longenecker K L, et al. The effect of ionic liquids on protein crystallization and X-ray diffraction resolution[J]. Crystal Growth & Design, 2009, 9(8): 3463-3469.
12	Gomes J M, Silva S S, Reis R L. Biocompatible ionic liquids: fundamental behaviours and applications[J]. Chemical Society Reviews, 2019, 48(15): 4317-4335.
13	Gao Y A, Zhao X Y, Dong B, et al. Inclusion complexes of beta-cyclodextrin with ionic liquid surfactants[J]. The Journal of Physical Chemistry B, 2006, 110(17): 8576-8581.
14	Wei Y, Wang F, Zhang Z Q, et al. Micellization and thermodynamic study of 1-alkyl-3-methylimidazolium tetrafluoroborate ionic liquids in aqueous solution[J]. Journal of Chemical & Engineering Data, 2014, 59(4): 1120-1129.
15	Cheng X H, Bai X Q, Jing S, et al. Self-assembly of imidazolium-based rodlike ionic liquid crystals: transition from lamellar to micellar organization[J]. Chemistry - A European Journal, 2010, 16(15): 4588-4601.
16	戴国亮, 于泳, 康琦, 等. 溶菌酶晶体生长前期溶液中聚集体研究[J]. 化学学报, 2004, 62(8): 757-761.
	Dai G L, Yu Y, Kang Q, et al. Study of the aggregates in lysozyme solution before crystal growth[J]. Acta Chimica Sinica, 2004, 62(8): 757-761.
17	李德海, 迟玉杰. 溶菌酶活力的简易测定[J]. 中国乳品工业, 2002(5): 128-129.
	Li D H, Chi Y J. Facility detection of lysozyme activity[J]. China Dairy Industry, 2002(5): 128-129.
18	Proteau A, Shi R, Cygler M. Application of dynamic light scattering in protein crystallization[J]. Current Protocols in Protein Science, 2010, 61(1): 17.10.1-17.10.9.
19	Mosquera V, Ruso J M, Prieto G, et al. Characterization of the interactions between lysozyme and n-alkyltrimethylammonium bromides by zeta potential measurements[J]. The Journal of Physical Chemistry, 1996, 100(41): 16749-16753.
20	McPherson A. Introduction to protein crystallization[J]. Methods, 2004, 34(3): 254-265.
21	王克夷. 疏水作用和蛋白质[J]. 生命的化学, 1999, 19(5): 233-235.
	Wang K Y. Hydrophobic interaction and protein [J].Chemistry of Life, 1999, 19(5): 233-235.
22	韩葵葵. 黄酮类药物与血清蛋白的相互作用以及不同表面活性剂的影响[D]. 扬州: 扬州大学, 2008.
	Han K K. The interaction between flavonoids and serum proteins and the influence of different surfactants [D].Yangzhou: Yangzhou University, 2008.
23	Walker A R, Baddam N, Cisneros G A. Unfolding pathways of hen egg-white lysozyme in ethanol[J]. The Journal of Physical Chemistry B, 2019, 123(15): 3267-3271.
24	Dang L P, Fang W Z, Li Y, et al. Ionic liquid-induced structural and activity changes in hen egg white lysozyme[J]. Applied Biochemistry and Biotechnology, 2013, 169(1): 290-300.
25	Chanphai P, Bekale L, Tajmir-Riahi H A. Effect of hydrophobicity on protein-protein interactions[J]. European Polymer Journal, 2015, 67: 224-231.
26	Sarmiento F, Ruso J M, Prieto G, et al. ζ-potential study on the interactions between lysozyme and sodium n-alkylsulfates[J]. Langmuir, 1998, 14(20): 5725-5729.
27	Guo Y, Zhang B, Lu C, et al. Locating the binding domains of lysozyme with ionic liquids in aqueous solution via spectroscopic studies[J]. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2019, 214: 239-245.
28	To B C S, Lenhoff A M. Hydrophobic interaction chromatography of proteins (Ⅲ): Transport and kinetic parameters in isocratic elution[J]. Journal of Chromatography A, 2008, 1205(1/2): 46-59.
29	Gao A T, Wu Q H, Wang D H, et al. A superhydrophobic surface templated by protein self-assembly and emerging application toward protein crystallization[J]. Advanced Materials, 2016, 28(3): 579-587.
30	Kwaambwa H M, Rennie A R. Interactions of surfactants with a water treatment protein from Moringa oleifera seeds in solution studied by zeta-potential and light scattering measurements[J]. Biopolymers, 2012, 97(4): 209-218.
31	Zhou R B, Cao H L, Zhang C Y, et al. A review on recent advances for nucleants and nucleation in protein crystallization[J]. CrystEngComm, 2017, 19(8): 1143-1155.
32	Frizzo C P, Gindri I M, Bender C R, et al. Effect on aggregation behavior of long-chain spacers of dicationic imidazolium-based ionic liquids in aqueous solution[J]. Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2015, 468: 285-294.
33	解莹, 戴国亮. 光散射技术在蛋白质晶体生长研究中的应用和进展[J]. 化学通报, 2005, 68(2): 94-99.
	Xie Y,Dai G L. The application and development of laser light scattering in the investigation of protein crystal growth[J]. Chemistry, 2005, 68(2): 94-99.
34	Nanev C. Recent insights into the crystallization process; protein crystal nucleation and growth peculiarities; processes in the presence of electric fields[J]. Crystals, 2017, 7(10): 310.
35	Asanov A N, Delucas L J, Oldham P B, et al. Interfacial aggregation of bovine serum albumin related to crystallization conditions studied by total internal reflection fluorescence[J]. Journal of Colloid and Interface Science, 1997, 196(1): 62-73.

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溴化1-辛基-3-甲基咪唑聚集状态对蛋白质结晶的影响研究

Study on the effect of 1-octyl-3-methylimidazole bromide aggregation state on protein crystallization

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