Fundamental Processes in the Folding of Helical Proteins

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Protein folding, the processes by which an initially unfolded protein rapidly self assembles into its folded functional form, has often been called the second half of the genetic code. All of the information required for a protein to fold is encoded in the primary sequence but a detailed understanding of the folding process is still lacking. Interest in protein folding continues to grow, driven in part by the realization of the fundamental and practical importance of folding and in part by experimental and theoretical advances. The potential for synergy between computational simulation and experiments has generated considerable interest in small proteins that fold very rapidly. These are excellent test cases for addressing fundamental issues in protein folding and for developing new methodology. This project aims to deduce general mechanistic features of the folding of alpha-helical proteins, to test specific hypotheses about their folding, to develop new methods to study their folding, and to provide rigorous bench marks for computational studies. A key goal is to link experimental results with computational studies of folding. To that end, the research is primarily focused on the villin headpiece helical subdomain (HP-36). HP-36 is the smallest naturally occurring sequence which folds cooperatively and it is one of the fastest folding proteins known. Its rapid folding, small size and simple structure have made it an extraordinarily popular model system for computational and theoretical studies of protein folding. Key questions to be addressed by this research include: (1) What is the nature of the unfolded state ensemble and what is its role in rapid folding? (2) Just how accurate are molecular dynamic simulations of protein folding, do they provide a reliable picture of the folding pathway or just a model of the native state? (3) What is the nature of the rate limiting step for folding? The unfolded state ensemble will be studied using peptide models, destabilized mutants, pH dependent stability measurements and simulations. Newly developed isotope edited IR detected T-jump methods will be used to study the folding of wild type HP-36 and mutants which alter the unfolded state. An improved understanding of protein folding has important implications for basic biology as well as important practical implications for biotechnology and for biomaterials design. The solution of the protein folding problem lies at the interface of chemistry, biology and physics and the research will involve close collaboration between experimental and theoretical groups. These factors create a stimulating multidisciplinary research environment for participating graduate and undergraduate students. Stony Brook is one of the most ethnically diverse major research universities in the nation and the research will enhance opportunities for traditionally underrepresented groups. Undergraduate participation is a key component of the research. Partnerships with undergraduate institutions have been established to expand undergraduate participation beyond Stony Brook. An "adopt a lab" program is being developed as part of the first year undergraduate honors lab class in order to provide a wide range of students with initial exposure to modern research. Traditional undergraduate laboratory exercises are being revised to better tie them into modern science. For example, certain food dyes are inhibitors of protein aggregation. Aggregation can be monitored using turbidity measurements conducted with inexpensive undergraduate spectrometers. The experiment illustrates light scattering, protein structure, the concept of inhibition and provides an introduction to protein folding while nicely tying into current research. Lab exercises will also be developed as part of Stony Brook's university-wide program designed to provide support and encouragement to undergraduate women who are interested in science and engineering.