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Changing the Apoptosis Pathway through Evolutionary Protein Design

Shultis, David, Mitra, Pralay, Huang, Xiaoqiang, Johnson, Jarrett, Khattak, Naureen Aslam, Gray, Felicia, Piper, Clint, Czajka, Jeff, Hansen, Logan, Wan, Bingbing, Chinnaswamy, Krishnapriya, Liu, Liu, Wang, Mi, Pan, Jingxi, Stuckey, Jeanne, Cierpicki, Tomasz, Borchers, Christoph H., Wang, Shaomeng, Lei, Ming, Zhang, Yang
Journal of molecular biology 2019 v.431 no.4 pp. 825-841
apoptosis, calorimetry, caspase-9, luminescence, mutation, peptides, proteins, proteolysis
One obstacle in de novo protein design is the vast sequence space that needs to be searched through to obtain functional proteins. We developed a new method using structural profiles created from evolutionarily related proteins to constrain the simulation search process, with functions specified by atomic-level ligand–protein binding interactions. The approach was applied to redesigning the BIR3 domain of the X-linked inhibitor of apoptosis protein (XIAP), whose primary function is to suppress the cell death by inhibiting caspase-9 activity; however, the function of the wild-type XIAP can be eliminated by the binding of Smac peptides. Isothermal calorimetry and luminescence assay reveal that the designed XIAP domains can bind strongly with the Smac peptides but do not significantly inhibit the caspase-9 proteolytic activity in vitro compared with the wild-type XIAP protein. Detailed mutation assay experiments suggest that the binding specificity in the designs is essentially determined by the interplay of structural profile and physical interactions, which demonstrates the potential to modify apoptosis pathways through computational design.