Molecular modeling studies are proposed to investigate the structural and thermodynamic effects of mutations on the leucine zipper, a recently discovered sequence motif that mediates dimerization in a large family of eukaryotic transcription factors. The long term objective of this work is to understand the underlying basis of the structure and stability of this dimerization motif. The proposed research will provide insights into structure-function relationships, molecular recognition, helix-packing, and protein stability that are particularly relevant to transcriptional regulation and oncogenesis. The specific goals of the proposed research are to calculate the effect on structure and stability of substitutions of both hydrophobic and polar residues at positions in the dimer interface and of modifications to salt bridges in the GCN4 leucine zipper, with particular emphasis on dissecting the energetics of stability changes into relative contributions due to the hydrophobic effect, packing interactions, helix-forming tendency, buried and surface hydrogen bonds, and other inter- and intramolecular interactions. Free energy computer simulations coupled with a component analysis procedure for analyzing the results will be employed, and results will be carefully compared with experimentally determined crystal structures and measurements of thermodynamic stability. Interpretations from the molecular modeling studies will be used to construct an understanding of the structure and stability of the leucine zipper. This understanding will be tested by proposing new mutations that will be studied experimentally and compared to the theoretical results.
