董晓燕
蛋白质复性和纯化;蛋白质稳定性;基因工程
个性化签名
- 姓名:董晓燕
- 目前身份:
- 担任导师情况:
- 学位:
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学术头衔:
博士生导师
- 职称:-
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学科领域:
生物化学工程
- 研究兴趣:蛋白质复性和纯化;蛋白质稳定性;基因工程
董晓燕,教授、博导
主要教育经历
1990年,日本信州大学农学部,农学硕士。
1991年,日本综研化学株式会社,研究员。
1996年,日本九州大学农学部,高级访问学者。
1997年,天津大学化工学院,副教授。
2002年,天津大学化工学院,教授。
2005年,天津大学化工学院,工学博士。
2007年,天津大学化工学院,博导。
主要研究方向
蛋白质复性和纯化
蛋白质稳定性
基因工程
主要学术成就、奖励及荣誉
中国高校自然科学奖二等奖(2002年,第二完成人);
天津市自然科学一等奖(2004年,第二完成人);
天津大学“三八”红旗手(2006年);
目前承担的主要科研项目
教育部回国人员资助基金;
教育部优秀青年教师基金;
国家“973”计划项目;
天津市自然基金2项(其中重点基金1项)
国家自然科学基金5项;
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主页访问
3174
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关注数
0
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成果阅读
1249
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成果数
18
【期刊论文】Refolding of denatured lysozyme assisted by artificial chaperones in reverse micelles
董晓燕, Xiao-Yan Dong, Xiao-Yue Wu, Yan Sun*
Biochemical Engineering Journal 31 (2006) 92-95,-0001,():
-1年11月30日
This article reports oxidative protein refolding assisted by artificial chaperones in reverse micelles formed by nonionic surfactant of sorbitan trioleate modified with Cibacron Blue F-3GA in n-hexane. The denatured/reduced lysozymewas used as a model protein and cetyltrimethylammonium bromide (CTAB) and-cyclodextrin as the artificial chaperones. The use of the artificial chaperones has proved to increase the refolding efficiency of denatured-reduced lysozyme at the concentration range studied (3.5-5.9 mg/mL). Moreover, the artificial chaperones increased the refolding yield in a wide range of urea concentrations. However, the optimal urea concentration range was little affected by the presence of the artificial chaperones.
Protein refolding, Lysozyme, Reverse micelles, Artificial chaperone, Nonionic surfactant, Cibacron Blue F-3GA
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引用
董晓燕, Yang Liu, Xiao-Yan Dong, Yan Sun*
Biochemical Engineering Journal 28 (2006) 281-288,-0001,():
-1年11月30日
Sorbitan trioleate was modified with Cibacron Blue F-3GA (CB) to create an affinity surfactant and to form affinity-based reverse micelles in nhexane. The partitioning equilibria and the extraction kinetics of lysozyme and bovine serum albumin (BSA) were then examined. The solubilization capacity of the reverse micellar system for lysozyme increased linearly with increasing the CB concentration from 0.1 to 0.5 mmol L−1. In contrast, the capacity for BSA at 0.5 mmol L-1 of coupled CB was only about one-fifth that for lysozyme. It indicates a strong steric hindrance effect of the micelles for the high molecular mass protein. The overall volumetric mass transfer coefficient of lysozyme in the forward extraction increased from 0.43×10−3 to 1.25×10−3 s−1 with increasing CB concentration from 0.1 to 0.5 mmol L−1. Due to the high molecular mass of BSA, its volumetric mass transfer coefficient in the forward extraction was only one-sixth that of lysozyme. The ratio of the coefficient in the back extraction to that in the forward extraction was less than 0.03, much lower than those in other micellar systems. It indicates that the interfacial resistance in this system was severer than in others.
Affinity, Reverse micelles, Nonionic surfactant, Protein, Equilibrium, Extraction kinetics
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董晓燕, Yang Liu, Xiao-Yan Dong, Yan Sun*
Journal of Colloid and Interface Science 297 (2006) 805-812,-0001,():
-1年11月30日
Sorbitan trioleate (Span 85) modified with Cibacron Blue F-3GA (CB) was used as an affinity surfactant (CB-Span 85) to form affinity-based reversed micelles in n-hexane. It was found that the addition of hexanol to the reversed micellar system resulted in a significant increase in water content and hydrodynamic radius of the affinity-based reversed micelles. Moreover, the reversed micelles with hexanol revealed broader aggregation number distribution and larger average aggregation number than the reversed micelles without hexanol addition. This is considered to be due to the decreases in the micellar curvature and rigidity of the micellar interfacial layer and the increase in the micellar interfacial fluidity. Consequently, the solubilization capacity of lysozyme increased about 70% in the reversed micellar solution with 3 vol% hexanol. On the other hand, the capacity of BSA was only 30% increased under the same conditions due to its larger molecular size than lysozyme. Kinetic analysis revealed that the increase in the micellar interfacial fluidity in the presence of hexanol resulted in faster release of lysozyme from the micelles, thus leading to an increase of the overall volumetric mass transfer coefficient in the back extraction.
Affinity-based reversed micelles, Span 85, Cibacron Blue F-3GA, Water content, Hydrodynamic radius, Solubilization, Proteins
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引用
董晓燕, 王玮, 白姝, 孙彦
化工学报,2006,57(1):74-78,-0001,():
-1年11月30日
利用多角度激光光散射仪和圆二色谱仪研究小分子水溶性一元醇和多元醇对溶菌酶分子的稳定性以及对变性溶菌酶的空间构象和折叠复性的影响。发现在一元醇和二元醇溶液中溶菌酶分子的稳定性均随着醇浓度的增大而提高,而变性酶分子在醇溶液中形成过多的α螺旋和β折叠等二级结构不利于酶的折叠复性;只有较低浓度的多元醇和甲醇有辅助变性溶菌酶复性的作用,其中10%甲醇、20%甘油和30%乙二醇有利于促进酶的复性。
醇, 溶菌酶, 复性, 分子相互作用, 空间结构
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董晓燕, 曹俊, 俞海青,
天津大学学报,2006,39(1):29-33,-0001,():
-1年11月30日
为了促使多肽亲和配基在实际分离研究中得到广泛应用,利用Fmoc固相合成法对从噬菌体展示肽库中筛选得到的与胰岛素具有高特异性亲和的七肽片段(HWWWPAS)进行了合成。在合成中发现,肽段序列中脯氨酸的存在会严重影响合成结果。云除脯氨酸后,目标产物的收率从0%提高到11.6%;同时,七肽序列中丙氨酸的存在也会影响合成效果。若将丙氨酸除去,合成收率会明显提高(达到76%),特别是将丙氨酸更换为谷氨酸产物的收率和线度可达到70%以上。利用固定化六肽片段(HWWWES)进行亲和吸附实验的结果表明,改造后的六肽配基与原先的七肽配基具有相似的亲和吸附效果。
多肽亲和配基, Fmoc固相合成, 氨基酸序列, 亲和吸附
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引用
董晓燕, Yuan Li, Xiao-Yan Dong, Yan Sun*
Biochemical Engineering Journal 27 (2005) 33-39,-0001,():
-1年11月30日
Interest in producing large quantities of plasmid DNA has recently increased as a result of the rapid evolution of gene therapy and DNA vaccines. Hydrophobic chromatography is a popular technique in the downstream processing of plasmid DNA. However, the low capacity and the high mass transfer resistance of most commercially available packings for bio-macromolecules limit their application in a large-scale process. In this work, a hydrophobic absorbent with wide pores was synthesized by the solid porogenic method. Analyses by scanning electron microscopy and mercury intrusion porosimetry revealed that the matrix contained two families of pores, i.e., micropores smaller than 100 nm and superpores of 500-7300 nm. The superpores provided not only convective flow channels for the mobile phase, but also a large surface for biomolecules binding. So the chromatographic process can be operated at high flow rate with high column efficiency and low backpressure as identified on a 2-mL column (5mm i.d., 2cm length). With a loading up to 2.6 mg of 5.4 kb plasmid (pcDNA3) in 8mL feedstock and operated at a flow rate as high as 20 cm/min, nearly 100% of plasmid was recovered with a purity of 100%. The results indicate that the hydrophobic medium is promising for high-speed purification of plasmid DNA.
Plasmid DNA, Hydrophobic interaction chromatography, Biporous medium, Purification, High speed
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董晓燕, 黄艳, 孙彦
天津大学学报,2004,37(7):643-646,-0001,():
-1年11月30日
研究了多种添加剂促进变性还原溶菌酶复性的作用。考察了添加剂浓度及变性剂盐酸胍浓度对复性收率的影响。结果表明,精氨酸、乙酰胺、丙酮、硫脲及甘油均能有效促进变性溶菌酶复性,并且存在最佳的添加剂浓度使变性溶菌酶的复性收率最大。在促进复性中,乙酰胺等结构类似物与盐酸胍具有相同作用,因此,在降低复性液中盐酸胍浓度的同时,适当提高乙酰胺浓度即可获得较高的复性收率。当盐酸胍浓度为0.2mol/L时,复性收率达到90%时的乙酰胺浓度为2mol/L,但降低盐酸胍浓度至0.06mol/L时,达到相同变性收率的乙酰胺浓度需要4mol/L。甘油与盐酸胍存在着协同作用,在一定浓度的盐酸胍存在下,添加适量的甘油能获得较高的复性收率。
溶菌酶, 复性, 稀释剂, 变性
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【期刊论文】流加操作与稀释添加耦合辅助高浓度变性蛋白质复性动力学
董晓燕, 史广泉, 李琳, 孙彦
化工学报,2004,55(8):1313-1318,-0001,():
-1年11月30日
研究了流加操作与稀释添加耦合辅助高浓度变性还原溶菌酶的复性,建立了相关的过程动力学模型。利用三态复性的过程模型对实验数据进行了拟合,获得了较好的拟合效果。利用模型分析了复性过程的动力学特性,结果表明,两者耦合可在较低的盐酸胍浓度( mol·L-1)存在下使较高浓度的(5mg·ml-1)变性溶菌酶获得80%以上的复性收率;并且发现添加剂乙酰胺的辅助复性作用与盐酸胍相同,降低盐酸胍浓度,适当提高乙酰胺浓度即可使聚集体生成速率常数最小,酶的复性收率最大;但甘油与盐酸胍具有协同作用,只有在适当的盐酸胍浓度下添加甘油才可获得理想的复性效果。
复性, 流加操作, 添加剂, 乙酰胺, 甘油, 盐酸胍, 溶菌酶
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董晓燕, Hai-Qing Yu, Xiao-Yan Dong, Yan Sun
Chromatographia 2004, 60, October (No.7/8),-0001,():
-1年11月30日
A heptapeptide phage display library was screened with insulin to find its ligands for affinity chromatography. The peptide was synthesized and coupled to EAH Sepharose 4B (5.4umol mL-1 bed). Then, insulin chromatography was carried out with mobile phases of different pH values and by the addition of urea and ethylene glycol. It was found that electrostatic interactions were predominant for the affinity binding, and hydrogen bonding might also contribute somewhat to the affinity. Finally, frontal analysis was performed and the dynamic binding capacity of the affinity column for insulin at 50% breakthrough was estimated at 60.6mg mL-1 bed, which was about two times higher than the theoretical binding capacity of the monomeric insulin. The result suggests that insulin was bound in dimer state in a stoichiometric relationship with the coupled peptide, indicating the high binding efficiency of the peptide ligand for insulin.
Column liquid chromatography Affinity chromatography Peptide ligand Phage display library Insulin
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【期刊论文】Refolding kinetics of denatured-reduced lysozyme in the presence of folding aids
董晓燕, Xiao-Yan Dong, Yan Huang, Yan Sun*
Journal of Biotechnology 114(2004)135-142,-0001,():
-1年11月30日
The refolding kinetic behavior of denatured-reduced lysozyme in the presence of folding aids (acetamide, acetone, thiourea, l-arginine or glycerol) was studied utilizing a simplified model describing the competition between first-order folding reaction and third-order aggregation. It was found that the protein folding aids could be categorized into two groups. One of them at proper concentrations, such as acetamide, acetone, thiourea and l-arginine, stabilized unfolded protein or folding intermediates. In the presence of these additives, the folding rate decreased with increasing their concentration, and there existed a concentration where the aggregation rate constant was minimized. So, there was an optimum concentration for the folding aids to produce a high yield. The other group was protein stabilizers such as glycerol. In the presence of this kind of folding aids, both the refolding rate and yield were enhanced by increasing their concentration to a proper value. Moreover, their effect on improving protein refolding was additive to those of the first group. So the cooperative application of the two kinds of folding aids could result in favorable refolding rate and yield of protein.
Protein refolding, Kinetics, Folding aids, Dilution additives, Model
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