Structure & Function of Bacteriophage Portal Proteins

University Of Alabama At Birmingham, Birmingham AL

Investigators

Abstract

Intellectual Merit The project focuses on deciphering the sequence of protein/protein interactions which occur during the assembly of viral capsids using the bacteriophage phi29 as a model system. Viral capsids are protective protein shells that self assemble from hundreds of chemically identical protein molecules. In the final capsid these molecules are precisely positioned in space with an overall spherical form. The protein shell surrounds and protects the viral nucleic acid and in one class of capsid the shell assembles first and the nucleic acid is subsequently pumped into the shell through a conduit known as a portal. Despite the ubiquity of this architectural theme, little is known about the pathway or sequence of protein/protein interaction through which the proteins self-assemble. Recent experimental data suggests that assembly nucleates from a complex composed of multiple copies of two proteins, a scaffolding protein and the conduit forming portal protein. The project will use chemical cross-linking/mass spectrometry, hydrogen/deuterium exchange studies and mutational analysis in concert with computational docking to determine the detailed structure of this nucleation complex, and use this information to chemically stabilize the nucleation complex. The stabilized nucleation complexes will then be used to seed assembly reactions both in bulk solution and in single molecule experiments to probe the sequence and kinetics of subunit addition to growing capsids and derive a molecular level understanding of the assembly pathway. Broader Impact A detailed quantitative description of the molecular pathway of viral capsid assembly is required by the cadre of physicist and mathematicians who are developing models of self-assembly using viruses as a paradigm and a description at this level of detail is not currently available for any virus. The research project itself employs a wide variety of biophysical and biochemical tools and serves as an ideal training platform for the graduate and undergraduate students who will be carrying out the experiments. Finally, because detailed molecular pathways are best appreciated and understood when illustrated by animation the assembly pathway defined by this project will be animated for a lay audience by students in the Department of Art and the Art History Department exposing them to frontline science while allowing them to refine their animation skills and develop a portfolio.