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Molecular characterization of CPS II and ACT genes form Trypanosoma cruzi

克氏锥虫 CPS II 和 ACT 基因的分子特征

基本信息

批准号：
07457068
负责人：
AOKI Takashi
金额：
$ 4.86万
依托单位：
Juntendo University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07457068/
关键词：
Trypanosoma pyrimidine synthesis CPS II ACT spliced leader (SL)single copy gene pulsed field gel electrophoresis gene mapping スプライスリーダー(SL)アスパラギン酸トランスカルバモイラーゼスプライストリーダー(SL)

项目摘要

The purpose of this study was to characterize the Trypanosoma cruzi genes encoding carbamoyl-phosphate synthetase II (CPS II) and aspartate carbamoyltransferase (ACT), the first and second enzymes of de novo pyrimidine nucleotide biosynthesis. De novo pyrimidine synthesis is essential in most of organisms and the initial three enzymes, CPS II,ACT,and hihydroorotase, are expressed as a multi-functional protein, CAD,in higher eukaryotes including mammals. On the other hand, we demonstrated that, in Trypanosoma cruzi, CPS II and ACT genes were encoded independently in its genome and localized simultaneously on two different chromosomes. As far as we know, there has been no report about a distribution of closely related genes and a transcriptional system of such genes in trypanosomal genome. From these views, we have examined the genomic structure of the CPS II and ACT genes in T.cruzi. A genomic library from the parasite was screened by using CPS II- and ACT-specific probes, and positive clones obtained were aligned so as to detect an overlapping region between them. The results showed that the CPS II gene was located at 10 kb upstream of the ACT gene. In addition, we have obtained two different restriction maps of these genes, corresponding to each allele and reflecting the plasticity of trypanosome genome. Interestingly, we have also found the ACT genes which are not linked to CPS II,suggesting the existence of homologue(s) or pseudogene(s) of ACT in T.cruzi.

本研究的目的是表征克氏锥虫基因编码氨甲酰磷酸合成酶II（CPS II）和天冬氨酸氨甲酰转移酶（ACT），从头嘧啶核苷酸生物合成的第一和第二个酶。嘧啶的从头合成在大多数生物体中是必需的，并且最初的三种酶CPS II、ACT和二氢乳清酸酶在高等真核生物包括哺乳动物中表达为多功能蛋白CAD。另一方面，我们证明，在克氏锥虫，CPS II和ACT基因编码独立的基因组和本地化同时在两个不同的染色体上。迄今为止，还没有关于锥虫基因组中密切相关基因的分布及其转录系统的报道。从这些观点来看，我们已经研究了CPS II和ACT基因的基因组结构的克氏锥虫。通过使用CPS II和ACT特异性探针筛选寄生虫的基因组文库，并将获得的阳性克隆进行比对，以检测它们之间的重叠区域。结果表明，CPS Ⅱ基因位于ACT基因上游10 kb处。此外，我们还获得了这些基因的两个不同的限制性酶切图谱，分别对应于每个等位基因，反映了锥虫基因组的可塑性。有趣的是，我们还发现了ACT基因不与CPS II，这表明存在的同源物（S）或假基因的ACT克氏锥虫。