Cloning, Purification, and Analysis of a Novel Aryl Hydrocarbon Receptor Deletion Construct
Poster Number
26
Introduction/Abstract
The aryl hydrocarbon receptor, AhR, is a bHLH/PAS family transcription factor that can be activated by xenobiotic ligand binding. The downstream targets of AhR are involved in the detoxification, exocytosis, and inhibition of chemical carcinogenesis. Cell growth, differentiation, proliferation, and apoptosis have been associated with AhR activity. Recent studies have focused on the role of AhR in cancer pathways due to its role in suppressing tumor initiation, promotion and progression. We created a deletion construct in order to study the properties and interactions of the transcriptional activation domain of AhR, AhR N∆515.
Purpose
To clone, purify, and analyze function of the AhR the deletion construct, AhR N∆515.
Method
Primers were designed to amplify the transcriptional activation domain of AhR, which was cloned into the His-tagged pQE-80L bacterial vector. The bacterial expressed protein was purified under denaturing conditions using cobalt affinity beads, and refolded via limited dialysis. The refolded protein was analyzed by testing protein-protein interactions to a known binding partner via Co-Immunoprecipitation.
Results
The final AhR N∆515 was confirmed to function similar to the transactivation domain of the full-length AhR.
Significance
We have many N-terminal constructs of AhR available at our disposal, but this is our first C-terminal construct. AhR N∆515 will allow us to analyze properties of the transactivation domain.
Location
DeRosa University Center, Stockton campus, University of the Pacific
Format
Poster Presentation
Cloning, Purification, and Analysis of a Novel Aryl Hydrocarbon Receptor Deletion Construct
DeRosa University Center, Stockton campus, University of the Pacific
The aryl hydrocarbon receptor, AhR, is a bHLH/PAS family transcription factor that can be activated by xenobiotic ligand binding. The downstream targets of AhR are involved in the detoxification, exocytosis, and inhibition of chemical carcinogenesis. Cell growth, differentiation, proliferation, and apoptosis have been associated with AhR activity. Recent studies have focused on the role of AhR in cancer pathways due to its role in suppressing tumor initiation, promotion and progression. We created a deletion construct in order to study the properties and interactions of the transcriptional activation domain of AhR, AhR N∆515.
