CHARACTERIZATION OF ASXL2 AS AN INTRINSICALLY DISORDERED PROTEIN

During development and differentiation, the transcription regulation plays an important rule. The Polycomb Group (PcG) and the Trithorax Group (TrxG) proteins are two protein families; they act by forming multiprotein complexes that regulate transcription by modifying chromatin configurations through histone methylation. The establishment or the maintenance mistakes of the correct chromatin configuration have been connected to several diseases, including leukemia and muscular dystrophy. The activities of PcG and TrxG proteins are regulated by a special group of proteins known as Enhancers of trithorax and Polycomb (ETP). The Additional sex combs-like 2 (Asxl2) protein, which is expressed from the gene called additional sex combs like 2 (Drosophila) (Asxl2), belongs to the Enhancer of trithorax and Polycomb group (ETP). The Asxl2 protein is highly expressed in heart during both development and in adult life. In a genetic study, the Asxl2-/- mice either died as neonates with congenital heart abnormalities or displayed progressive deterioration of ventricular function. Although Asxl2 protein is important, there have been no significant studies of the Asxl2 protein’s structure. Protein functions are determined to a considerable degree by structure, so the structural studies will provide valuable information on Asxl2. Failures of protein folding into native structure generally produce inactive or toxic proteins. Misfolded proteins are believed to be the causes of some neurodegenerative diseases, including Alzheimer’s and Parkinson's diseases. ix Through investigation in silico, it is known that the Asxl2 protein contains a large native unstructured region, and this result might indicate the Asxl2 is an intrinsically disordered protein (IDP). Most IDPs are known to have regulatory roles. Owing to their lack of folded structure, the intrinsically disordered proteins display a flexible and random-coil-like conformation under physiological conditions, and have several unique features. This suggests that distinctive methods will be required to characterize their structure. In order to understand the structure of Asxl2, we have expressed and purified an Asxl2 N-terminal truncation that contains a large native unstructured region of two important domains: the HARE-HTH domain and the Asx homology domain (ASXH). We have then used Circular Dichroism (CD) as a means of investigating the structure of Asxl2. The CD spectra have been measured over a wavelength range from 250 nm to 190 nm at various temperatures, with and without Asxl2’s physiological binding partner, the ubiquity carboxyl-terminal hydrolase BAP1 (Bap1) protein in order to test the hypothesis that Asxl2 is an IDP.M.S. in Molecular Biochemistry and Biophysics, December 201