Computer-Aided Design of Thermostable Proteins

摘要

We are developing methods for computer-aided protein design and are testing these strategies by constructing thermostable variants of the lambda repressor. Repressor's DNA-binding domain normally denatures at 54C, and we have constructed a quadruple mutant that is stable to 71C and binds DNA as well as the wild type protein. Our fundamental goal is to develop methods for de nova protein design, and we are proceeding by treating the problem of protein design as an inverted version of the protein folding problem (Pabo, 1983). In protein folding, one is given an amino acid sequence and must predict how this folds in three dimensions. Protein design can be approached in quite a different way - one can begin by choosing a folded arrangement of the polypeptide backbone and then try to pick an amino acid sequence that will stabilize this structure. Inversion eliminates the problem of predicting long-range interactions, since residues which will interact in the final tertiary or quaternary structure already are close in space when they are added to the prefolded backbone. One should be able to pick residues which will have favorable interactions with their neighbors. We are developing a program, called PDB PROTEUS, for computer-aided protein design. Our program uses simple geometric aspects of protein structure and frequently uses local coordinate systems so that the geometric relationships are easier to visualize (Pabo and Suchanek, 1986). (aw)