Folding of a Multidomain Ribosomal Protein

Suny At Stony Brook, Stony Brook NY

Investigators

Abstract

0079406 Raleigh Elucidating how the amino acid sequence determines structure, the protein folding problem, is a central issue in modern structural biology. Recent developments have made this problem particularly timely. Massive genome sequencing efforts have lead to an explosion in our knowledge of the primary sequence of proteins. Unfortunately it is still impossible to predict structure from sequence. In principle, a more thorough understanding of the folding process should aid efforts to decipher the code that links sequence and structure. A detailed understanding of the folding process will also aid efforts to rationally modify proteins to enhance desired properties or confer new functions. The research described in this project focuses on experimental investigations of several issues in protein folding. The hypothesis that chain topology is the major determinant of folding kinetics will be examined by conducting studies of the folding of two sets of proteins. The folding kinetics of the C-terminal domain of the protein L9 will be measured and compared to theoretical predictions. In the intact protein, this domain appears to fold much more slowly than predicted by theory. The folding of the isolated domain will be studied in order to test if the deviation is caused by interactions with the rest of the protein or is more fundamental. The structure of the N-terminal domain of RNase HI is very similar to the N-terminal domain of L9 (NTL9) and the folding of this protein will be compared to NTL9. The role of the unfolded state in protein folding and stability will be examined using NTL9 as a model system. Each of the six acidic residues in NTL9 with be mutated and the pH dependence of the stability determined. These measurements provide information about electrostatic interactions in the unfolded state. The formation of electrostatic interactions during the folding process will be studied by conducting kinetic measurements with this set of mutants. These studies will be complimented by experiments that probe the development of the hydrophobic core of NTL9 during folding. Taken together, the kinetic studies will provide a detailed picture of the folding of NTL9.