2005 Retreat: Zheng-yu Peng Folding, Stability, and Function of Ankyrin Repeat Proteins

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Our laboratory studies the folding, stability, protein-protein interaction, and function of ankyrin repeat proteins. Ankyrin repeat is a 33-residue sequence motif that has been identified in hundreds of proteins throughout both prokaryotic and eukaryotic genomes. Most ankyrin repeat proteins are mediators of specific protein-protein interactions. Each ankyrin repeat folds into two anti-parallel a-helices and a b-hairpin. Part of the inner a-helix and the b-hairpin is used in molecular recognition and protein-protein interactions. We have previously designed a "consensus" ankyrin repeat protein and determined the minimal folding unit of ankyrin repeat is a two-repeat module. In this talk, I will focus on two new research projects recently completed in the lab. First, we have analyzed the effect of individual surface residues on the interaction between GABPa and GABPb using alanine scanning mutagenesis and isothermal titration calorimetry. The result showed that a hydrophobic "hot spot" that is responsible for most of the interaction free energy. Second, we have analyzed the effect of several stabilizing substitutions on the wild type and mutant tumor suppressor p16 using a mammalian cell line. We are able to show that these substitutions significantly reduce the level of aggregation of mutant p16 proteins in vivo and promote kinase assembly by immunofluorescence analysis. The result is consistent with the measure of cell cycle arrest by flow cytometry.