Hydrophobin (HFBI):: A potential fusion partner for one-step purification of recombinant proteins from insect cells
Hydrophobin (HFBI):: A potential fusion partner for one-step purification of recombinant proteins from insect cells
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DOI：
10.1016/j.pep.2007.12.014
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发表时间：
2008-05-01
影响因子：
1.6
通讯作者：
Oker-Blom, Christian
中科院分区：
文献类型：
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作者：
Lahtinen, Tomi;Linder, Markus B.;Oker-Blom, Christian
疏水蛋白在真菌表面结构的结合和组装以及介质-空气相互作用中起着重要作用。当涉及大分子的纯化时，这些疏水性质提供了有趣的可能性。在基于表面活性剂的水胶束两相系统中，水溶性疏水蛋白浓缩在胶束结构内，并因此分布到限定的水相。当涉及重组蛋白的大规模生产时，这种一步纯化特别有吸引力。在本研究中，疏水蛋白HFBI的里氏木霉表达作为一个N-末端融合鸡亲和素在杆状病毒感染的昆虫细胞。通过共聚焦显微镜分析重组融合构建体的细胞内分布，随后通过使用非离子表面活性剂在水胶束两相系统中从细胞质提取物纯化蛋白质。结果表明，疏水蛋白和其亲和素融合体在昆虫细胞中有效表达，并且这些疏水蛋白可以通过采用含水胶束两相系统在一个步骤中从这些细胞中有效纯化。(C)2008年爱思唯尔公司All rights reserved.
Hydrophobins play an important role in binding and assembly of fungal surface structures as well as in medium-air interactions. These, hydrophobic properties provide interesting possibilities when purification of macromolecules is concerned. In aqueous micellar two-phase systems, based on surfactants, the water soluble hydrophobins are concentrated inside micellar structures and, thus, distributed to defined aqueous phases. This, one-step purification is attractive particularly when large-scale production of recombinant proteins is concerned. In the present study the hydrophobin HFBI of Trichoderma reesei was expressed as an N-terminal fusion with chicken avidin in baculovirus infected insect cells. The intracellular distribution of the recombinant fusion construct was analyzed by confocal microscopy and the protein subsequently purified from cytoplasmic extracts in an aqueous micellar two-phase system by using a non-ionic surfactant. The results show that hydrophobin and an avidin fusion thereof were efficiently expressed in insect cells and that these hydrophobic proteins could be efficiently purified from these cells in one-step by adopting an aqueous micellar two-phase system. (C) 2008 Elsevier Inc. All rights reserved.