ROLE OF CHAPERONES IN MAINTAINING THE ASPARAGINE REPEAT-RICH PROTEOME OF P. FALCI

描述（由申请人提供）： 摘要 PI：MURALIDHARAN，VASANT 项目：1 K99 AI 099156 -01标题：伴侣蛋白在维持P. peculci天冬酰胺重复丰富蛋白质组中的作用登录号：3400178 ================== 注意事项：本摘要摘自应用程序，未经SRA验证。当应用程序扫描过程出现问题时，提取的文本可能不正确或不完整。 ================== 恶性疟原虫是一种致命的人类病原体，每年造成100多万人死亡。恶性疟原虫的蛋白质组不成比例地富含天冬酰胺重复序列;四分之一的蛋白质含有这样的重复序列。蛋白质中Asn重复序列的存在导致淀粉样原纤维的形成。在恶性疟原虫的生命周期中，它经历了大的温度波动，这可以促进淀粉样纤维的形成。分子伴侣或热休克蛋白，如Hsp 110、Hsp 70和Hsp 40，存在于所有生命界并影响淀粉样纤维的形成。然而，它们的生物学作用和作用机制仍不明确。拟议的职业发展计划的目的是获得基本的见解伴侣在维护的Asn重复丰富的蛋白质组的恶性疟原虫的功能，同时建立一个独立的学术生涯在大学环境。候选人将在生物化学、蛋白质生物物理学和寄生虫学的坚实基础上，在疟疾寄生虫红细胞内阶段研究的先驱和领导者丹尼尔戈德堡博士的指导下，发展成为恶性疟原虫生物学的独立研究人员。该计划将在华盛顿大学医学院传染病系医学系实施，该学院是生物医学科学的领先机构。在最近的一篇论文中，该候选人报告了一种新型可调控的荧光亲和（RFA）标签的开发，该标签用于研究恶性疟原虫蛋白酶体亚基Rpn 6中的28个残基Asn重复序列。Asn重复序列不影响蛋白质的功能，稳定性或相互作用，即使在热休克下，这表明存在一个积极的过程，确保正确折叠的Asn重复含有蛋白质。在指导阶段，候选人将：1）使用PfHsp 110基因已通过生化和显微镜方法进行RFA标记的寄生虫系，检查PfHsp 110在寄生虫蛋白质稳定中的功能。淀粉样蛋白形成蛋白也将在这些寄生虫系中表达，以研究PfHsp 110如何影响原纤维形成。2)通过用PfHsp 110突变体和其他生物体的同源物补充RFA标记的寄生虫，确定作用机制和特异性。PfHsp 110也将在异源哺乳动物系统中表达，并将使用生物化学和显微镜方法研究其对哺乳动物细胞中原纤维形成的影响。在奖项的独立阶段，候选人将阐明寄生虫的Hsp 70，Hsp 70样和Hsp 40分子伴侣在维持稳定的Asn重复丰富的蛋白质组中的作用。这些分子伴侣被发现的候选人与PfHSP 110。研究计划中概述的生物化学，显微镜和细胞生物学方法的培训使候选人能够作为独立研究人员开展全面而富有成效的研究计划。拟议的研究将揭示恶性疟原虫伴侣蛋白功能的基本方面，这将为疟疾以及其他蛋白质错误折叠疾病提供新的药物靶点。相关性疟疾是一种致命疾病，困扰着全世界数亿人。导致最致命形式疟疾的寄生虫恶性疟原虫正在迅速获得抗药性，因此必须确定杀死这种寄生虫的新策略。拟议的研究将调查新类别的药物靶标，以了解它们在寄生虫生物学中的作用，并开发更好的药物。

DESCRIPTION (provided by applicant): Abstract PI: MURALIDHARAN, VASANT Project: 1K99AI099156-01 Title: ROLE OF CHAPERONES IN MAINTAINING THE ASPARAGINE REPEAT-RICH PROTEOME OF P. FALCI Accession Number: 3400178 ================== NOTICE: THIS ABSTRACT WAS EXTRACTED FROM APPLICATION AND HAS NOT BEEN PROOFED BY AN SRA.WHEN THERE ARE PROBLEMS WITH THE APPLICATION SCANNING PROCESS, THE EXTRACTED TEXT MAY BE INCORRECT OR INCOMPLETE. ================== Plasmodium falciparum is a deadly human pathogen that is responsible for over a million deaths every year. The proteome of P. falciparum is disproportionately rich in repeats of asparagine; one in four proteins contain such repeats. The presence of Asn repeats in a protein leads to amyloid fibril formation. During its life cycle, P. falciparum undergoes large temperature fluctuations that can promote amyloid fibril formation. Chaperones or heat shock proteins, such as Hsp110, Hsp70 and Hsp40, are present in all kingdoms of life and influence amyloid fibril formation. However, their biological roles and mechanism of action remains poorly defined. The proposed career development plan aims to gain fundamental insights into the function of chaperones in maintaining the Asn repeat-rich proteome of P. falciparum, while establishing an independent academic career in a university setting. The candidate will build on his strong foundation in biochemistry, protein biophysics and parasitology to develop into an independent researcher in P. falciparum biology under the mentorship of Dr. Daniel Goldberg, a pioneer and leader in the study of the intraerythrocytic stages of the malaria parasite. The plan will be carrid out in the Department of Medicine, Division of Infectious Diseases at Washington University School of Medicine, a leading institution in biomedical science. In a recent paper, the candidate reported the development of a novel regulatable, fluorescent, affinity (RFA) tag that was used to study a 28-residue Asn repeat in the P. falciparum proteasome subunit, Rpn6. The Asn repeat did not affect the protein function, stability or interactions even under heat shock; suggesting th presence of an active process that ensures proper folding of Asn repeat containing proteins. During the mentored phase, the candidate will: 1) Examine the function of PfHsp110 in parasite proteostasis using the parasite lines where PfHsp110 gene has been RFA tagged via biochemical and microscopic approaches. Amyloid forming proteins will also be expressed in these parasite lines to study how the PfHsp110 influences fibril formation. 2) Determine the mechanism of action and specificity by complementing the RFA tagged parasite with mutants of PfHsp110 and homologs from other organisms. PfHsp110 will also be expressed in a heterologous mammalian system and its effect on fibril formation in mammalian cells will be studied using biochemical and microscopic approaches. In the independent phase of the award the candidate will elucidate the roles of Hsp70, Hsp70-like and Hsp40 chaperones of the parasite in maintaining a stable Asn repeat-rich proteome. These chaperones were discovered by the candidate to be associated with PfHsp110. Training in biochemical, microscopic and cell biological approaches as outlined in the research plan has equipped the candidate to embark on a comprehensive and fruitful research program as an independent researcher. The proposed studies will reveal fundamental aspects of chaperone function in P. falciparum, which will furnish new drug targets against malaria as well as other protein misfolding diseases. Relevance Malaria is a deadly disease that afflicts hundreds of millions of people worldwide. The parasite that causes the deadliest form of malaria, Plasmodium falciparum, is rapidly gaining drug resistance making it imperative to identify new strategies to kill this parasite. The proposed research will investigate new classes of drug targets to understand their role in parasite biology and develop better drugs against them.