STRUCTURAL BASIS FOR ROTAVIRUS ENTRY INTO CELLS

轮状病毒胃肠炎是#年儿科疾病的主要原因。 发达国家的儿童死亡率和发展中国家的儿童死亡率。 轮状病毒是一种无包膜的二十面体病毒，它通过 对膜的直接穿透过程知之甚少。一种理解 这一过程将有助于开发预防和治疗 针对轮状病毒的措施，并可能为预防轮状病毒提供新的策略 治疗药物的细胞内靶向。轮状病毒外壳 蛋白质，VP4和VP7，在进入过程中脱落，很可能是膜的中介物 穿透力。进入需要切割尖峰蛋白VP4，这是一种 血凝素、毒力和中和决定因素，以及可能的细胞 附着蛋白。杆状病毒表达的VP4将被提纯。 由胰酶裂解引起的构象变化将通过 酶敏感性、凝胶过滤、圆二向色性和脂质体 颠覆。VP4将被结晶，并具有高分辨率的结构 决心将会被启动。外衣壳糖蛋白VP7， 经历一种钙依赖的构象变化，与非 涂层，是一个中和决定因素，并与VP4和 膜。VP7将被提纯，结构研究分析其 将进行构象变化和与VP4的相互作用。 我是哈佛大学传染病综合培训的研究员 程序。我建议研究轮状病毒进入细胞的结构基础 在斯蒂芬·哈里森博士的实验室里，对一个 病毒的数量正在进行中。我在美国完成了医学博士课程 斯坦福大学从事轮状病毒的抗原性和分子研究。我的 对临床重要感染的科学方法感兴趣 疾病的灵感来自扎伊尔和巴基斯坦的工作，而生物 哈佛大学人类学专业的学生。我近期的职业目标是 接受生物化学和结构生物学方面的培训以补充 在病毒学、免疫学和分子生物学方面的培训 研究生院。我的长期职业目标是应用基础科学 临床重要病毒学问题的技术。

Rotavirus gastroenteritis is a major cause of pediatric illness in developed countries and of childhood mortality in developing countries. Rotavirus is a non-enveloped icosahedral virus that enters cells by the poorly understood process of direct membrane penetration. An understanding of this process would aid in developing preventive and therapeutic measures against rotavirus and might suggest novel strategies for the intracellular targeting of therapeutic agents. The rotavirus outer capsid proteins, VP4 and VP7, are shed during entry and likely mediate membrane penetration. Entry requires cleavage of the spike protein VP4, which is a hemagglutinin, virulence and neutralization determinant, and probable cell attachment protein. Baculovirus-expressed VP4 will be purified. Conformation changed induced by trypsin cleavage will be assayed by protease sensitivity, gel filtration, circular dichroism, and liposome disruption. VP4 will be crystallized, and a high resolution structural determination will be initiated. VP7, the outer capsid glycoprotein, undergoes a calcium-dependent conformation change associated with un- coating, is a neutralization determinant, and interacts with VP4 and membranes. VP7 will be purified, and structural studies analyzing its conformation changes and interaction with VP4 will be undertaken. I am a fellow in the Harvard Combined Infectious Diseases Training Program. I propose to study the structural basis for rotavirus cell entry in Dr. Stephen Harrison's laboratory, where structural research on a number of viruses is ongoing. I completed an M.D.-Ph.D. program at Stanford, pursuing antigenic and molecular studies on rotavirus. My interest in scientific approaches to clinically important infectious diseases was inspired by work in Zaire and Pakistan while a biological anthropology student at Harvard College. My immediate career goal is to acquire training in biochemistry and structural biology to complement training in virology, immunology, and molecular biology obtained in graduate school. My long term career goal is to apply basic scientific techniques to clinically important virologic problems.