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绿猴多瘤病毒1型VP1蛋白C端区域在病毒粒子形成中的作用。

Role of the C-terminal region of vervet monkey polyomavirus 1 VP1 in virion formation.

作者信息

Yamaguchi Hiroki, Kobayashi Shintaro, Maruyama Junki, Sasaki Michihito, Takada Ayato, Kimura Takashi, Sawa Hirofumi, Orba Yasuko

机构信息

Division of Molecular Pathobiology, Research Center for Zoonosis Control, Hokkaido University, Sapporo, Hokkaido 001-0020, Japan.

出版信息

J Vet Med Sci. 2014 May;76(5):637-44. doi: 10.1292/jvms.13-0568. Epub 2014 Jan 13.

Abstract

Recently, we detected novel vervet monkey polyomavirus 1 (VmPyV) in a vervet monkey. Among amino acid sequences of major capsid protein VP1s of other polyomaviruses, VmPyV VP1 is the longest with additional amino acid residues in the C-terminal region. To examine the role of VmPyV VP1 in virion formation, we generated virus-like particles (VLPs) of VmPyV VP1, because VLP is a useful tool for the investigation of the morphological characters of polyomavirus virions. After the full-length VmPyV VP1 was subcloned into a mammalian expression plasmid, the plasmid was transfected into human embryonic kidney 293T (HEK293T) cells. Thereafter, VmPyV VLPs were purified from the cell lysates of the transfected cells via sucrose gradient sedimentation. Electron microscopic analyses revealed that VmPyV VP1 forms VLPs with a diameter of approximately 50 nm that are exclusively localized in cell nuclei. Furthermore, we generated VLPs consisting of the deletion mutant VmPyV VP1 (ΔC VP1) lacking the C-terminal 116 amino acid residues and compared its VLP formation efficiency and morphology to those of VLPs from wild-type VmPyV VP1 (WT VP1). WT and ΔC VP1 VLPs were similar in size, but the number of ΔC VP1 VLPs was much lower than that of WT VP1 VLPs in VP1-expressing HEK293T cells. These results suggest that the length of VP1 is unrelated to virion morphology; however, the C-terminal region of VmPyV VP1 affects the efficiency of its VLP formation.

摘要

最近,我们在一只绿猴体内检测到了新型绿猴多瘤病毒1型(VmPyV)。在其他多瘤病毒主要衣壳蛋白VP1的氨基酸序列中,VmPyV VP1最长,其C末端区域有额外的氨基酸残基。为了研究VmPyV VP1在病毒粒子形成中的作用,我们制备了VmPyV VP1的病毒样颗粒(VLP),因为VLP是研究多瘤病毒病毒粒子形态特征的有用工具。将全长VmPyV VP1亚克隆到哺乳动物表达质粒中后,将该质粒转染到人胚肾293T(HEK293T)细胞中。此后,通过蔗糖梯度沉降从转染细胞的裂解物中纯化出VmPyV VLP。电子显微镜分析显示,VmPyV VP1形成直径约50 nm的VLP,且仅定位于细胞核中。此外,我们制备了由缺失C末端116个氨基酸残基的缺失突变体VmPyV VP1(ΔC VP1)组成的VLP,并将其VLP形成效率和形态与野生型VmPyV VP1(WT VP1)的VLP进行了比较。WT和ΔC VP1 VLP大小相似,但在表达VP1的HEK293T细胞中,ΔC VP1 VLP的数量远低于WT VP1 VLP。这些结果表明,VP1的长度与病毒粒子形态无关;然而,VmPyV VP1的C末端区域会影响其VLP形成的效率。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/c91e/4073331/38f43102ec64/jvms-76-637-g001.jpg

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