BACKGROUND Malaria parasites form intracellular membranes that separate the parasite from the internal space of erythrocytes, and membrane proteins from the parasites are exported to the host via the membrane. In our previous study, Plasmodium vivax early transcribed membrane protein (PvETRAMP) 11.2, an intracellular membrane protein that is highly expressed in blood-stage parasites, was characterized as a highly immunogenic protein in P. vivax malaria patients. However, the other PvETRAMP family proteins have not yet been investigated. In this study, PvETRAMPs were expressed and evaluated to determine their immunological profiles. METHODS The protein structure and amino acid alignment were carried out using bioinformatics analysis software. A total of six PvETRAMP family proteins were successfully expressed and purified using a wheat germ cell free protein expression system and the purified proteins were used for protein microarray and immunization of mice. The localization of the protein was determined with serum against PvETRAMP4. IgG subclasses were assessed from the immunized mice. RESULTS In silico analysis showed that P. vivax exhibits nine genes encoding the ETRAMP family. The ETRAMP family proteins are relatively small molecules with conserved structural features. A total of 6 recombinant ETRAMP proteins were successfully expressed and purified. The serum positivity of P. vivax malaria patients and healthy individuals was evaluated using a protein microarray method. Among the PvETRAMPs, ETRAMP4 showed the highest positivity rate of 62%, comparable to that of PvETRAMP11.2, which served as the positive control, and a typical export pattern of PvETRAMP4 was observed in the P. vivax parasite. The assessment of IgG subclasses in mice immunized with PvETRAMP4 showed high levels of IgG1 and IgG2b. PvETRAMP family proteins were identified and characterized as serological markers. CONCLUSIONS The relatively high antibody responses to PvETRAMP4 as well as the specific IgG subclasses observed in immunized mice suggest that the ETRAMP family is immunogenic in pathogens and can be used as a protein marker and for vaccine development.

中文翻译：

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评估间日疟原虫对早期转录膜蛋白家族的抗体反应。

背景技术疟原虫形成将寄生虫与红细胞的内部空间分开的细胞内膜，并且来自寄生虫的膜蛋白通过膜输出至宿主。在我们之前的研究中，间日疟原虫早期转录膜蛋白（PvETRAMP）11.2是一种在血液阶段寄生虫中高表达的细胞内膜蛋白，在间日疟原虫疟疾患者中被表征为高度免疫原性蛋白。但是，尚未研究其他PvETRAMP家族蛋白。在这项研究中，表达并评估了PvETRAMPs的免疫学特征。方法使用生物信息学分析软件进行蛋白质结构和氨基酸比对。使用无小麦生殖细胞的蛋白质表达系统成功表达和纯化了总共六种PvETRAMP家族蛋白质，纯化的蛋白质用于蛋白质微阵列和小鼠免疫。用抗PvETRAMP4的血清确定蛋白质的定位。从免疫小鼠评估IgG亚类。结果在计算机分析中，间日疟原虫显示9个编码ETRAMP家族的基因。ETRAMP家族蛋白是具有保守结构特征的相对较小的分子。总共成功表达和纯化了6种重组ETRAMP蛋白。使用蛋白质微阵列方法评估间日疟原虫疟疾患者和健康个体的血清阳性。在PvETRAMP中，ETRAMP4的阳性率最高，为62％，与PvETRAMP11.2相当，用作阳性对照，并且在间日疟原虫寄生虫中观察到典型的PvETRAMP4输出模式。对用PvETRAMP4免疫的小鼠中IgG亚类的评估显示出高水平的IgG1和IgG2b。鉴定了PvETRAMP家族蛋白并将其表征为血清学标志物。结论在免疫小鼠中观察到的对PvETRAMP4的相对较高的抗体应答以及特定的IgG亚类表明ETRAMP家族在病原体中具有免疫原性，可以用作蛋白质标记物并用于疫苗开发。鉴定了PvETRAMP家族蛋白并将其表征为血清学标志物。结论在免疫小鼠中观察到的对PvETRAMP4的相对较高的抗体应答以及特定的IgG亚类表明ETRAMP家族在病原体中具有免疫原性，可以用作蛋白质标记物并用于疫苗开发。鉴定了PvETRAMP家族蛋白并将其表征为血清学标志物。结论在免疫小鼠中观察到的对PvETRAMP4的相对较高的抗体应答以及特定的IgG亚类表明ETRAMP家族在病原体中具有免疫原性，可以用作蛋白质标记物并用于疫苗开发。