Molecular dynamics of the H-ras gene-encoded p21 protein; identification of flexible regions and possible effector domains.
We previously reported a complete computer-based three-dimensional structure for residues 1-171 of the Gly 12-containing ras-gene-encoded p21 protein complexed with GDP. This structure was subsequently shown to closely agree with a high-resolution x-ray crystallographic structure of p21. In this communication, we report a molecular dynamics stimulation of the modelled structure in an explicit shell of water molecules to identify domains within the protein that are unusually flexible. These domains represent regions which are most likely to undergo important conformational changes when the protein is activated by binding to GTP or by oncogenic amino acid substitutions such as Val for Gly 12. The starting structure was surrounded with water molecules, temperature-equilibrated and then followed over a 100 ps trajectory during which time the energy converged after about 50 ps. Regions of the protein that were found to have the largest coordinate fluctuations involved residues 12-16, 30-35, 40-52, 60-73, 85-89, 101-109, 119-123, and 127-131. Many of these sequences with high flexibility have been implicated in the functioning of this protein. Since the overall largest fluctuations were observed for residues 101-106 and 119-123, p21 peptides containing these residues (96-110 and 115-126) were synthesized and were found to inhibit strongly the effects of oncogenic p21 protein in an oocyte maturation assay. These results indicate that the flexible p21 sequences may constitute critical functional domains of the activated protein and that this general approach may be useful for identification of important functional domains in proteins.