CAREER: Conformations of Beta-Sheet Folding Intermediates: The Discrimination Between Productive Folding and Aggregation

University Of Notre Dame, Notre Dame IN

Investigators

Abstract

The objective of this CAREER project is to develop general models for b-sheet formation and stabilization. Several all-b-sheet proteins are unusually thermostable, and many function in harsh extracellular environments where unusually high stability is required for activity. Many of these proteins have simple structural topologies (for example, the parallel b-helix), which, together with their thermostability, make them attractive subjects for use as 'molecular scaffolds' for designing new functionalities. However, these proteins have an Achilles' heel: while the native protein may be thermostable, the folding intermediates are often quite thermolabile, and prone to aggregation. A prerequisite for adapting these proteins for alternative functions is to understand how they operate. Yet little is known about the details of b-sheet formation, particularly parallel b-sheet formation. This project will use biochemical and biophysical approaches such as fluorescence spectroscopy, protease sensitivity, and changes in molecular radius to examine the conformations of folding intermediates for two all-b-sheet proteins with very stable yet difficult-to-form native states. This work will uncover the features of b-sheet folding intermediates that (1) promote correct folding to the native structure, and (2) suppress the off-pathway aggregation reaction. The results of these studies will have broad application to the design of correctly folded b-sheet proteins for biotechnological purposes. The education goals of this project focus on introducing students at all levels to the interdisciplinary approaches of modern scientific research and development. The specific objectives include updating the undergraduate curriculum to incorporate a new upper-level undergraduate elective in protein biophysics, and a new general chemistry course for first year students, focused on the interdisciplinary approaches of modern scientific discovery. Current efforts to recruit female and minority researchers will be continued and expanded.