Molecular expression of biologically active substances produced from newborn larvae of Trichinella

旋毛虫新生幼虫产生的生物活性物质的分子表达

基本信息

批准号：
12670229
负责人：
TAKAHASHI Yuzo
金额：
$ 1.09万
依托单位：
Gifu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

项目摘要

Trichinella have a number of biologically active substances, some of which may be endowed with unique characteristics and possible usage for experimental and pharmaceutical purposes. We have already cloned and expressed a serine proteinase inhibitor of T. spiralis of muscle larvae. Mass production of serine proteinase inhibitor by genetic engineering may provide a new means for pharmaceutical usage and vaccine.Newborn larvae penetrate the host tissue and migrate through the body of the host. As a result of immunoscreening of the cDNA library constructed from not only muscle larvae but also newborn larvae, we could clone a novel excretory-secretory (E-S) protein, myosin, tropomyosin and serine proteinase. The recombinant proteins were produced in an Escherichia coli expression system. The recombinant protein of serine proteinase and enolase exhibited enzymatic activity. The enzymatic activity of recombinant serine proteinase could be mainly plasmin-like proteinase activity.We developed a detection system for myogenic regulatory factors such as MyoD and myogenin. MyoD and myogenin were expressed from the early phase of cystogenesis in Trichinella infection.We investigated gene expression at different developmental stages of Trichinella. The analysis genes included E-S proteins and macrophage migration inhibitory factor. There were many differences in the expression level of these proteins at different developmental stages.After this, we would like to search more useful proteins from Trichinella cDNA library, and study the function of these proteins by using the recombinant proteins expressed in insect cells.

旋毛虫具有许多生物活性物质，其中一些可能被赋予独特的特性并可能用于实验和制药目的。我们已经克隆并表达了肌肉幼虫螺旋毛虫的丝氨酸蛋白酶抑制剂。利用基因工程大规模生产丝氨酸蛋白酶抑制剂，可能为医药用途和疫苗提供新的手段。新生幼虫可穿透宿主组织，在宿主体内迁移。通过对肌肉幼虫和新生幼虫构建的 cDNA 文库进行免疫筛选，我们可以克隆一种新的排泄分泌 (E-S) 蛋白：肌球蛋白、原肌球蛋白和丝氨酸蛋白酶。重组蛋白在大肠杆菌表达系统中产生。丝氨酸蛋白酶和烯醇化酶的重组蛋白表现出酶活性。重组丝氨酸蛋白酶的酶活性主要是纤溶酶样蛋白酶活性。我们开发了MyoD和肌生成素等生肌调节因子的检测系统。 MyoD和myogenin在旋毛虫感染的囊肿发生早期就开始表达。我们研究了旋毛虫不同发育阶段的基因表达。分析基因包括E-S蛋白和巨噬细胞迁移抑制因子。这些蛋白在不同发育阶段的表达水平存在很大差异。此后，我们希望从旋毛虫cDNA文库中寻找更多有用的蛋白，并利用在昆虫细胞中表达的重组蛋白来研究这些蛋白的功能。