Identification of the major cysteine protease of Giardia and its role in encystation
Identification of the major cysteine protease of Giardia and its role in encystation
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DOI：
10.1074/jbc.m802133200
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发表时间：
2008-06-27
影响因子：
4.8
通讯作者：
Sajid, Mohammed
中科院分区：
文献类型：
--
作者：
DuBois, Kelly N.;Abodeely, Marla;Sajid, Mohammed
蓝氏贾第鞭毛虫是一种原生动物寄生虫，也是真核生物最早的分支。贾第鞭毛虫的生命周期在无性繁殖的营养型和感染性包囊型之间交替。包囊和脱囊是贾第虫生存和传播的关键过程。贾第虫体内的半胱氨酸蛋白酶参与蛋白水解过程，使生命周期在包囊和脱囊过程中得以延续。使用实时定量PCR，在营养生长和成囊期间测量贾第虫基因组中27个家族CA半胱氨酸蛋白酶基因的表达。贾第虫半胱氨酸蛋白酶2是最高度表达的半胱氨酸蛋白酶在两个生命周期阶段的测量，与一个显着的表达增加，在包囊。贾第虫半胱氨酸蛋白酶2的mRNA转录在成囊期间上调7倍，并且比任何其他贾第虫蛋白酶基因产物高3倍以上。重组贾第虫半胱氨酸蛋白酶2的表达，纯化和生化特性。重组半胱氨酸蛋白酶2蛋白的活性被证实与G. Lamblia裂解物。贾第虫半胱氨酸蛋白酶2与包囊特异性囊泡中的包囊壁蛋白在包囊形成过程中共定位，并将包囊壁蛋白2加工成贾第虫包囊壁中发现的大小。这些数据表明，贾第鞭毛虫半胱氨酸蛋白酶2不仅是主要的半胱氨酸内切蛋白酶表达的贾第鞭毛虫，但也是中央的成囊过程。
Giardia lamblia is a protozoan parasite and the earliest branching clade of eukaryota. The Giardia life cycle alternates between an asexually replicating vegetative form and an infectious cyst form. Encystation and excystation are crucial processes for the survival and transmission of Giardia. Cysteine proteases in Giardia have been implicated in proteolytic processing events that enable the continuance of the life cycle throughout encystation and excystation. Using quantitative real-time PCR, the expression of twenty-seven clan CA cysteine protease genes in the Giardia genome was measured during both vegetative growth and encystation. Giardia cysteine protease 2 was the most highly expressed cysteine protease during both life cycle stages measured, with a dramatic expression increase during encystation. The mRNA transcript for Giardia cysteine protease 2 was 7-fold up-regulated during encystation and was greater than 3-fold higher than any other Giardia protease gene product. Recombinant Giardia cysteine protease 2 was expressed, purified, and biochemically characterized. The activity of the recombinant cysteine protease 2 protein was confirmed to be identical to the dominant cysteine protease activity found in G. lamblia lysates. Giardia cysteine protease 2 was co-localized with cyst wall protein in encystation-specific vesicles during encystation and processed cyst wall protein 2 to the size found in Giardia cyst walls. These data suggest that Giardia cysteine protease 2 is not only the major cysteine endoprotease expressed in Giardia, but is also central to the encystation process.