EXPRESSION AND PURIFICATION OF RECOMBINANT ASXL2 PROTEINS

Polycomb Group (PcG) and the Trithorax Group (TrxG) proteins are two highly conserved proteins families which regulate transcription by modifying chromatin structure. The activity of these two groups of proteins is regulated by a special group of proteins known as Enhancers of Trithorax and Polycomb (ETP). Recently our lab has generated a mutant mouse model for an ETP Group gene, Additional sex combs-like 2 (Asxl2) (Baskind 2009). Asxl2 is highly expressed in heart during development and in adult life. Depending on the genetic background Asxl2 -/- mice either die as neonates with congenital heart conditions or live to adulthood but develop heart dysfunction. In order to understand the structural basis of Asxl2’s function we propose to express and purify various truncations/deletions of Asxl2 that are important for its function. We expressed four different truncations and one deletion of Asxl2 in E.coli as GST fusion proteins. GST-Asxl2 fusions were purified using glutathione beads. This completes an important milestone in our structural study. In the future the purified recombinant proteins will be used for protein pull down assays and biophysical assays aimed at understanding the structure of Asxl2, which will improve knowledge of chromatin-based transcriptional regulation in the heart and will facilitate the detection, prevention and treatment of heart diseases.M.S. in Biology, July 201