Structural Basis for Rotavirus Entry into Cells

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.