To develop monovalent dengue virus-like particle for serotype 3 (DENV-LP/3), we prepared and expressed two structural polyprotein constructs using silkworm and Bm5 cells: DENV-3 Capsid-premembrane-envelope (DENV-3CprME) and premembrane-envelope (DENV-3prME). The expressed PA-tagged 3CprME and 3prME polypeptides were partially purified by PA-tag affinity chromatography and had molecular weights of 85 and 75 kDa, respectively. Expressed proteins were separately verified using the following primary antibodies: the anti-PA tag antibody, DENV premembrane polyclonal antibody, and DENV envelope polyclonal antibody. Transmission electron microscopy revealed that these DENV-3CprME and 3prME formed rough, spherical DENV-LPs (DENV-LP/3CprME and DENV-LP/3prME), respectively, with a diameter of 30–55 nm. The heparin-binding assay demonstrated that these DENV-LPs contained the envelope protein domain III on their surfaces. Both DENV-LPs showed an affinity to sera from human dengue patients and immunized mice. Immunization of mice with DENV-LP/3prME significantly induced the level of antibodies compared with DENV-LP/3CprME. These results indicate that DENV-LP/3prME is suitable as a vaccine candidate compared with DENV-LP/3CprME.

中文翻译：

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家蚕幼虫中登革热病毒样颗粒血清型3的产生及其引起小鼠体液免疫反应的能力。

要开发血清型3的单价登革热病毒样颗粒（DENV-LP / 3），我们使用蚕和Bm5细胞制备并表达了两种结构多蛋白构建体：DENV-3衣壳前膜包膜（DENV-3CprME）和前膜包膜（DENV-3prME）。表达的带有PA标签的3CprME和3prME多肽通过PA标签亲和色谱法部分纯化，分子量分别为85和75 kDa。使用以下主要抗体分别验证了表达的蛋白质：抗PA标签抗体，DENV前膜多克隆抗体和DENV包膜多克隆抗体。透射电子显微镜显示，这些DENV-3CprME和3prME分别形成直径为30-55 nm的粗糙球形球形DENV-LP（DENV-LP / 3CprME和DENV-LP / 3prME）。肝素结合测定表明这些DENV-LP在其表面上包含包膜蛋白结构域III。两种DENV-LP均显示出对人类登革热患者和免疫小鼠血清的亲和力。与DENV-LP / 3CprME相比，用DENV-LP / 3prME免疫小鼠可显着诱导抗体水平。这些结果表明，与DENV-LP / 3CprME相比，DENV-LP / 3prME适合作为候选疫苗。