Investigations of Binding Targets of the Pro-Mutagen 2-Aminoanthracene

Environmental exposures causing ingestions of toxic chemicals, such as the polycyclic aromatic hydrocarbon 2-aminoanthracene (2-AA), may increase the risk of developing cancer and other diseases such as diabetes. Some cytogical, histological, and immunocytochemical studies in the past have indicated that the endocrine and exocrine pancreas cellular morphology can be altered by chronic dietary exposure to 2-AA and can in turn cause diabetic-like symptoms. The goal of this study is to investigate the toxicity and mechanisms associated with the binding targets of 2-AA.

A new technique known as dynamic isoelectric anisotropy ligand binding assay (DIABLA) utilizes capillary isoelectric focusing and fluorescence anisotropy to separate proteins in their active form and evaluate chemical interactions. A high-voltage power supply is used at intervals along the capillary to control the shape of the electric field within the capillary, the pH gradient, and the location and width of the focused protein bands in the sample. Once focused, areas of non-zero anisotropy will be observed to determine where 2-AA is interacting with a protein. The protein may then be collected for further analysis by mass spectrometry since the focused band once migrated will remain at a designated sampling point. This will allow for the investigation of 2-AA binding as a step towards understanding the biological activities and effects through screening a complex mixture of proteins. DIABLA will be utilized to focus proteomic samples of human pancreas with 2-AA in an attempt to recognize and isolate the protein targets. This will lead to further understanding of 2-AA mode of action.