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新型穿山甲型重复蛋白 PfATRP 的分子特征和免疫反应模式

Molecular Characterization and Immuno-Reactivity Patterns of a Novel Armadillo-Type Repeat Protein, PfATRP.

机构信息

West African Center for Cell Biology of Infectious Pathogens, University of Ghana, Accra, Ghana.

Department of Biochemistry, Kogi State University, Anyigba, Nigeria.

出版信息

Front Cell Infect Microbiol. 2020 Mar 20;10:114. doi: 10.3389/fcimb.2020.00114. eCollection 2020.

DOI:10.3389/fcimb.2020.00114
PMID:32266165
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC7100384/
Abstract

Nearly half of the genes in the genome have not yet been functionally investigated. We used homology-based structural modeling to identify multiple copies of Armadillo repeats within one uncharacterized gene expressed during the intraerythrocytic stages, PF3D7_0410600, subsequently referred to as Armadillo-Type Repeat Protein (PfATRP). Soluble recombinant PfATRP was expressed in a bacterial expression system, purified to apparent homogeneity and the identity of the recombinant PfATRP was confirmed by mass spectrometry. Affinity-purified α-PfATRP rabbit antibodies specifically recognized the recombinant protein. Immunofluorescence assays revealed that α-PfATRP rabbit antibodies reacted with schizonts. Anti-PfATRP antibody exhibited peripheral staining patterns around the merozoites. Given the localization of PfATRP in merozoites, we tested for an egress phenotype during schizont arrest assays and demonstrated that native PfATRP is inaccessible on the surface of merozoites in intact schizonts. Dual immunofluorescence assays with markers for the inner membrane complex (IMC) and microtubules suggest partial colocalization in both asexual and sexual stage parasites. Using the soluble recombinant PfATRP in a screen of plasma samples revealed that malaria-infected children have naturally acquired PfATRP-specific antibodies.

摘要

基因组中的近一半基因尚未进行功能研究。我们使用基于同源性的结构建模方法，在一个在红细胞内阶段表达的未被表征的基因（PF3D7_0410600）中鉴定了多个 Armadillo 重复序列的拷贝，随后将其称为 Armadillo 重复蛋白（PfATRP）。可溶性重组 PfATRP 在细菌表达系统中表达，经纯化达到明显均一性，并通过质谱法确认重组 PfATRP 的身份。亲和纯化的α-PfATRP 兔抗体特异性识别重组蛋白。免疫荧光分析显示α-PfATRP 兔抗体与裂殖子反应。抗-PfATRP 抗体表现出围绕裂殖子的外周染色模式。鉴于 PfATRP 在裂殖子中的定位，我们在裂殖体阻滞试验中测试了其逸出表型，并证明在完整裂殖体中，天然 PfATRP 无法在裂殖子表面获得。与内膜复合物（IMC）和微管的标记物进行双重免疫荧光分析表明，在无性和有性阶段寄生虫中存在部分共定位。在血浆样本的筛选中使用可溶性重组 PfATRP 表明，感染疟疾的儿童已经产生了天然的 PfATRP 特异性抗体。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/f6b6564a30e7/fcimb-10-00114-g0007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/850ffff4107d/fcimb-10-00114-g0001.jpg
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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/3e0450bc2812/fcimb-10-00114-g0004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/1992d97eca9f/fcimb-10-00114-g0005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/76e3f3d6b572/fcimb-10-00114-g0006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/f6b6564a30e7/fcimb-10-00114-g0007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/850ffff4107d/fcimb-10-00114-g0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/10bd084d9dc0/fcimb-10-00114-g0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/ace087295ef2/fcimb-10-00114-g0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/3e0450bc2812/fcimb-10-00114-g0004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/1992d97eca9f/fcimb-10-00114-g0005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/76e3f3d6b572/fcimb-10-00114-g0006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/641b/7100384/f6b6564a30e7/fcimb-10-00114-g0007.jpg

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