• Purification of Recombinant Human Histone Methyltransferases for Inhibition Studies

      Jahan, Asmat; Shaikh, Zahid; Department of Chemistry & Physics (2017-03)
      Resistance to cellular apoptotic pathways is a major contributing factor in the metastasis of cancer. One such cell-intrinsic pathway which induces programmed cell death is ligand cell surface death receptors, including Fas protein. In primary colorectal cancer, Fas expression is often diminished. The silencing of Fas expression is perhaps a mechanism by which human colorectal cancer cells evade apoptosis. The decrease in Fas expression levels is associated with hypermethylation of histone 3 lysine 9 catalyzed by histone methyltransferases (HMTases) such as SUV39H1 and SUV39H2. Because of the role of methylation in silencing Fas expression, HMTase inhibitors represent potential anti-cancer agents. Verticillin A, a natural compound extracted from wild mushroom, is a broad-based HMTase inhibitor and has been shown to cause toxicity in mice. Our goal in this study is to identify specific SUV39H1 and SUV39H2 inhibitors that are less toxic than Verticillin A. To investigate this, human SUV39H1 was sub-cloned into an expression vector, transformed into an E.coli expression strain and purified using affinity chromatography. Gel electrophoresis and Western blot analysis confirmed the presence of partially purified human SUV39H1. The next phase of the project will involve testing the activity of the purified protein in an in vitro assay.