Chromatographically-purified capsid proteins of red-spotted grouper nervous necrosis virus expressed in Saccharomyces cerevisiae form virus-like particles
Chromatographically-purified capsid proteins of red-spotted grouper nervous necrosis virus expressed in Saccharomyces cerevisiae form virus-like particles
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DOI：
10.1016/j.pep.2013.03.007
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发表时间：
2013-06-01
影响因子：
1.6
通讯作者：
Kim, Hong-Jin
中科院分区：
文献类型：
--
作者：
Choi, Yoo Ri;Kim, Hyoung Jin;Kim, Hong-Jin
神经坏死病毒(NNV)导致海水鱼类大量死亡，给水产养殖户造成巨大的经济损失。预防NNV感染的候选疫苗是NNV病毒样颗粒(VLP)，它是由重组NNV衣壳蛋白组装而成的结构。NNV衣壳蛋白已在昆虫细胞和大肠杆菌表达系统中表达，并通过蔗糖和氯化铯密度梯度超速离心法纯化。在本研究中，我们在酿酒酵母(S.cerevisiae)中表达了赤点石斑鱼NNV(RGNNV)衣壳蛋白，并建立了一种具有大规模疫苗生产潜力的层析方法。用一步肝素层析成功纯化了RGNNV衣壳蛋白。透射电子显微镜证实纯化的RGNNV衣壳蛋白质量较高，呈VLP形式，平均直径25 nm，悬浮液均匀，无任何聚集现象。此外，RGNNV衣壳蛋白还能诱导小鼠产生抗RGNNV衣壳蛋白抗体。我们认为，RGNNV衣壳蛋白在酿酒酵母中表达，并经肝素层析纯化，具有足够的质量用于疫苗。(C)2013 Elsevier Inc.保留所有权利。
Nervous necrosis viruses (NNVs) cause mass mortality of marine fish, leading to large economic losses for aquaculturists. A promising vaccine candidate for preventing NNV infection is the NNV virus-like particle (VLP), which is a structure resulting from assembly of recombinant NNV capsid protein. NNV capsid proteins have been expressed in insect cells and the Escherichia coli expression system, and purified by non-scalable protocols such as ultracentrifugation on sucrose and cesium chloride density gradients. In this study, we expressed red-spotted grouper NNV (RGNNV) capsid proteins in Saccharomyces cerevisiae (S. cerevisiae) and developed a chromatography-based method with potential for large-scale vaccine production. The RGNNV capsid protein was successfully purified by a single-step of heparin chromatography. Transmission electron microscopy (TEM) confirmed the high quality of the purified RGNNV capsid protein: it was in the form of VLPs with mean diameters of 25 nm, in homogeneous suspension without any aggregation. Moreover, the RGNNV capsid protein elicited anti-RGNNV capsid protein antibodies in mice. We suggest that RGNNV capsid protein expressed in S. cerevisiae and purified by heparin chromatography, is of sufficient quality for use as a vaccine. (c) 2013 Elsevier Inc. All rights reserved.