Brydges S D, Sherman G D, Nockemann S, Loyens A, Däubener W, Dubremetz J F, Carruthers V B
The W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins University School of Hygiene and Public Health, 615 N. Wolfe Street, Baltimore, MD 21205, USA.
Mol Biochem Parasitol. 2000 Nov;111(1):51-66. doi: 10.1016/s0166-6851(00)00296-6.
During invasion of host cells, Toxoplasma gondii discharges the contents of small, apically located secretory organelles called micronemes. Micronemal proteins are known to be necessary for both parasite motility and invasion of host cells. To further define the contents of Toxoplasma micronemes, we used cell fractionation and secretion-modulating drugs to identify six novel, putative micronemal proteins. In this paper we describe preliminary characterization of one of these novel proteins, TgMIC5. Molecular cloning and DNA sequence analysis of the TgMIC5 cDNA and gene revealed that it encodes a previously identified immunodominant antigen called H4. TgMIC5 also possesses a consensus sequence unique to members of the parvulin family of peptidyl-prolyl cis-trans isomerases (PPIases). TgMIC5 is expressed as a preproprotein, which is proteolytically processed to a proprotein by signal peptidase before being further processed to a mature protein of 22 kDa. Using a combination of protein secretion experiments, immunofluorescence and immunoelectron microscopy, we demonstrated that TgMIC2 is stored in the micronemes of T. gondii tachyzoites before it is secreted into the surrounding medium. Based on its homology with parvulin-like PPIases, TgMIC5 may assist in the folding of other micronemal proteins that function in invasion of host cells by T. gondii tachyzoites.
在入侵宿主细胞的过程中，弓形虫会释放位于顶端的称为微线体的小分泌细胞器的内容物。已知微线体蛋白对于寄生虫的运动性和宿主细胞的入侵都是必需的。为了进一步确定弓形虫微线体的内容物，我们使用细胞分级分离和分泌调节药物来鉴定六种新的、假定的微线体蛋白。在本文中，我们描述了其中一种新蛋白TgMIC5的初步特征。TgMIC5 cDNA和基因的分子克隆及DNA序列分析表明，它编码一种先前鉴定的称为H4的免疫显性抗原。TgMIC5还具有肽基脯氨酰顺反异构酶（PPIases）的小菌素家族成员所特有的共有序列。TgMIC5表达为前体蛋白，在被进一步加工成22 kDa的成熟蛋白之前，先由信号肽酶蛋白水解加工成前体蛋白。通过结合蛋白质分泌实验、免疫荧光和免疫电子显微镜，我们证明了TgMIC2在被分泌到周围培养基中之前储存在弓形虫速殖子的微线体中。基于其与小菌素样PPIases的同源性，TgMIC5可能有助于弓形虫速殖子入侵宿主细胞过程中发挥作用的其他微线体蛋白的折叠。
