Investigating the Specificity of Coiled-Coil Recognition

The bZIP transcription factors make up a family of long α-helical proteins that dimerize based on a pattern of hydrophobic residues and bind to DNA through a region of basic residues. Because binding specificity is a particular topic of interest, the dimerization interaction is attractive as a possible candidate to better understand protein quaternary structure. Use of the Knob-Socket (KS) model for determination of packing structure provides a novel approach to analyze protein-protein interactions. A KS analysis of the protein-protein interface provides unique insight into the specificity of the classical leucine zipper pseudo-7mer repeat. From an analysis of the KS packing maps, this research provides evidence of a general framework for defining the specificity between coiled-coils. The KS maps show how hydrophobic specificity is defined in the coiled-coil interface, where knobs are centralized in the middle of the socket packing, while the peripheral socket residues are hydrophilic. Based on this KS analysis, the KS model will be used to design proteins that mimic the leucine zipper region of bZIP proteins. The proteins will be purified into E. coli and its 2º structure will be confirmed through circular dichroism. Binding specificity will be studied through mutations of the designed proteins and compared using the BACTH (bacterial adenylate cyclase two-hybrid) system