Title

Investigating the Specificity of Coiled Coil Recognition

Poster Number

8

Lead Author Affiliation

Pharmaceutical and Chemical Sciences/Chemistry

Lead Author Status

Masters Student

Second Author Affiliation

Pharmaceutical and Chemical Sciences/Chemistry

Second Author Status

Doctoral Student

Third Author Affiliation

Pharmaceutical and Chemical Sciences/Chemistry

Introduction

bZIP transcription factors make up a family of long α-helical proteins that dimerize and bind to DNA through a basic region that contains hydrophobic residues. They function as gene expression regulating factors and are therefore attractive possible candidates for small molecule binding. Binding specificity is a particular topic of interest. The development of an accurate method to characterize and map out the binding specificity of these bZIP proteins, would enable us to alter and possibly inhibit the function of its protein interactions.

Use of the Knob-Socket model for determination of packing structure provides a novel approach to analyze protein-protein as well as protein-nucleic acid interactions. A Knob-Socket analysis of the protein-protein interface provides unique insight into the classical leucine zipper pseudo-7mer repeat. A deeper analysis of longer leucine zippers shows unique packing patterns not indicated by classical representations like the helical wheel. From analysis of the Knob-Socket packing maps, this research provides evidence of a general framework for defining the specificity between coiled coils. the Knob-Socket 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. Furthermore, the bias of the filled over free propensities shows a clear pattern that explains the specificity of a set of hydrophobic interactions.

Purpose

The development of an accurate method to characterize and map out the binding specificity of these bZIP proteins, would enable us to alter and possibly inhibit the function of its protein interactions.

Method

The Knob-Socket 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 secondary 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.

Results

Secondary structure of the novel zipper sequences are confirmed through circular dichroism. Zipper constructs for the BACTH assay have been made and are tested for homo- and heterodimerization. Mutations to the zipper sequences, to test interactions of the coiled coils, have been designed.

Significance

Binding specificity between coiled coils still remains a question in the biochemistry field. The mapping of bZIP proteins, has not been done in a way to understand the different patterns that are found between the coiled coils. Mapping of known bZIP proteins using the Knob-Socket model allows one to see patterns in coiled coils. These patterns provide insight to how the binding specificity between coiled coils may be occurring. Through the use of mutations to novel bZIP-like proteins, specificity can be altered. By controlling the specificity of a protein this would get us one step closer to solving the question of binding specificity.

Location

DeRosa University Center

Format

Poster Presentation

Poster Session

Afternoon

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Apr 27th, 12:30 PM Apr 27th, 2:30 PM

Investigating the Specificity of Coiled Coil Recognition

DeRosa University Center

bZIP transcription factors make up a family of long α-helical proteins that dimerize and bind to DNA through a basic region that contains hydrophobic residues. They function as gene expression regulating factors and are therefore attractive possible candidates for small molecule binding. Binding specificity is a particular topic of interest. The development of an accurate method to characterize and map out the binding specificity of these bZIP proteins, would enable us to alter and possibly inhibit the function of its protein interactions.

Use of the Knob-Socket model for determination of packing structure provides a novel approach to analyze protein-protein as well as protein-nucleic acid interactions. A Knob-Socket analysis of the protein-protein interface provides unique insight into the classical leucine zipper pseudo-7mer repeat. A deeper analysis of longer leucine zippers shows unique packing patterns not indicated by classical representations like the helical wheel. From analysis of the Knob-Socket packing maps, this research provides evidence of a general framework for defining the specificity between coiled coils. the Knob-Socket 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. Furthermore, the bias of the filled over free propensities shows a clear pattern that explains the specificity of a set of hydrophobic interactions.