CHARACTERAZATION OF DBL DOMAIN PROTEINS IN P FALCIPARUM

恶性疟原虫中 DBL 结构域蛋白的表征

• 批准号: 6362344
• 负责人: DAVID Scott PETERSON
• 金额: $ 15.13万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-03-01 至 2003-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6362344
• 关键词: Plasmodium falciparum; SDS polyacrylamide gel electrophoresis; confocal scanning microscopy; electron microscopy; erythrocytes; fluorescence microscopy; gene targeting; genetic polymorphism; host organism interaction; immunoelectron microscopy; intracellular parasitism; laboratory rabbit; laboratory rat; northern blottings; polymerase chain reaction; protein binding; protein structure function; receptor binding; receptor expression; restriction fragment length polymorphism; single strand conformation polymorphism; southern blotting; tissue /cell culture; western blottings

Many of the specific molecular interactions that occur between the malarial parasite and its host, including cytoadherence, rosetting, and erythrocyte invasion, are mediated by parasite ligands encoded by proteins of the DBL-domain (Duffy binding like) superfamily. The DBL- domain is a cysteine rich domain with both conserved and variable features, and has been demonstrated to encode the ligand for the erythrocyte receptors glycophorin-A and the Duffy antigen. In the process of erythrocyte invasion P. falciparum is capable of utilizing several distinct pathways involving at least three different erythrocyte receptors. The only P. falciparum ligand known to mediate invasion by one of these pathways is EBA-175, a DBL-domain protein that binds to glycophorin-A. The hypothesis to be tested in this proposal is that several newly identified proteins of the DBL-domain family also function as ligands for erythrocyte receptors during invasion. The characterization of these proteins will involve determining their location within the parasite by fluorescence, electron and confocal microscopy and testing the ability of these proteins to bind erythrocytes. Preliminary investigation of the identity of the host receptor for these ligands will also be conducted, utilizing the binding regions expressed in a heterologous system, and assaying their ability to bind enzymatically modified erythrocytes, or those genetically deficient in defined surface molecules. The potential of using knockout parasites lacking the genes for these putative erythrocyte binding proteins will also be investigated. The extent to polymorphism within the genes encoding proteins shown to play a role in invasion will be investigated to suggest whether these proteins may be adapting to immune pressure or receptor heterogeneity. Finally, conserved features within the DBL-domains of the erythrocyte binding proteins will be exploited to identify additional members of this gene family which may also be involved in invasion.

许多特定的分子相互作用发生在 疟疾寄生虫及其宿主，包括细胞粘附，玫瑰花结， 红细胞侵入，是由寄生虫配体介导的， DBL结构域（Duffy binding like）超家族的蛋白质。 DBL- 结构域是一个富含半胱氨酸的结构域， 特征，并已被证明编码的配体的 红细胞受体血型糖蛋白A和达菲抗原。 在 红细胞侵入过程恶性疟原虫能够利用 涉及至少三种不同红细胞的几种不同途径 受体。 恶性疟原虫配体是唯一已知的介导 其中一个途径是EBA-175，它是一种DBL结构域蛋白， 血型糖蛋白-A。 本提案中要检验的假设是， DBL结构域家族的几种新鉴定的蛋白质也起作用， 作为红细胞受体的配体。 的 这些蛋白质的表征将涉及确定它们的 通过荧光、电子和共聚焦在寄生虫内定位 显微镜和测试这些蛋白质的结合能力， 红细胞 对宿主身份的初步调查 这些配体的受体也将进行，利用结合 在异源系统中表达的区域，并测定它们的能力 结合酶修饰的红细胞，或那些基因修饰的红细胞， 缺乏明确的表面分子。 使用knockout的潜力 缺乏这些假定的红细胞结合基因的寄生虫 还将研究蛋白质。 内的多态性程度 编码蛋白质的基因在入侵中起作用， 研究表明这些蛋白质是否可以适应免疫 压力或受体异质性。 最后，保存的功能， 红细胞结合蛋白的DBL结构域将被利用 为了鉴定该基因家族的其他成员， 参与入侵。