STRUCTURAL BASIS FOR ROTAVIRUS ENTRY INTO CELLS

轮状病毒胃肠炎是小儿疾病的主要原因， 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， 对直接膜渗透的过程知之甚少。的理解 这一过程将有助于发展预防和治疗 针对轮状病毒的措施，并可能提出新的战略， 治疗剂的细胞内靶向。轮状病毒外壳 蛋白质VP 4和VP 7在进入过程中脱落，并可能介导膜 渗透。进入需要刺突蛋白VP 4的切割，这是一种 血凝素、毒力和中和决定簇以及可能的细胞 附着蛋白将纯化杆状病毒表达的VP 4。 胰蛋白酶裂解诱导的构象变化将通过 蛋白酶敏感性、凝胶过滤、圆二色谱和脂质体 破坏VP 4将被结晶，并且高分辨率结构 将开始确定。VP 7，外壳糖蛋白， 经历了钙依赖的构象变化与非- 包被，是中和决定簇，与VP 4相互作用， 膜。将纯化VP 7，并对其进行结构研究分析。 将进行构象变化和与VP 4的相互作用。 我是哈佛传染病联合培训的研究员 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， 病毒数量正在增加。我完成了医学博士学位-博士项目 斯坦福大学，从事轮状病毒抗原和分子研究。我 对临床重要传染病的科学方法感兴趣 疾病的灵感来自于扎伊尔和巴基斯坦的工作，而生物 哈佛学院人类学专业的学生。我的职业目标是 获得生物化学和结构生物学方面的培训， 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 临床上重要的病毒学问题。

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.