Browsing by Subject "virtual screening"

The aims of this work were (1) to compare the three dimensional structures of different S- adenosylmethionine (SAM)-dependent methyltrasferases and (2) to screen in silico a commercial library for potential methyltransferase inhibitors. In this work we decided to focus on DNA methyltransferase-like enzyme (DNMT2) and catechol-O-methyltransferase(COMT). There were two different parts in my work. The first part was to analyze the 3Dstructures of DNMT2 and COMT in relation with their amino acid sequences. The structures of DNMT2 and COMT were compared together by means of superimposition with Sybyl 8. The ligand binding properties were studied by manual and automatic docking of known inhibitors in order to understand the binding specificity of these methyltransferases. The softwares I used for docking were Autodock 4.2 and Gold 4.0. The sequence alignments and superimposition of the known crystal structures showed that the structures of DNMT2 and COMT share a similar fold. Furthermore the main similarities between the structures of these enzymes are in the co-enzyme binding sites. The only significant difference in the binding sites is the place of one tyrosine residue, which causes a slight change in the conformation of the bound co-enzyme. Unlike co- enzyme binding sites, the substrate binding sites of DNMT2 and COMT are different. There is indeed a bound magnesium ion in the substrate binding site of COMT but not in the substrate binding site of DNMT2. Because the substrate binding sites are more different than the co-enzyme binding sites, we decided to screen the potential active ligands only at the substrate binding sites. The second part of the work was virtual screening. I used a subset of 20.000 molecules of ChemBridge DIVER Set that can be purchased commercially. The softwares I used for library preparation were CONCORD and Balloon, from which Balloon created more reasonable 3D structures for the docking. I did two parallel screenings to the crystal structure of COMT (PDB code 3BWM) with docking program GOLD 4.0, which is the only program that can take account metal coordination. To DNMT2 I did two sets of screenings, one with GOLD 4.0 and another with Autodock 4.2. I used known COMT inhibitors as control in the COMT run and known DNA methyltransferase inhibitors as control in DNMT2 run. Before docking to the three dimensional structure of DNMT2, one loop near the substrate binding site had to be modeled. I used Swiss-Modeler and Modeller softwares for that. Docking to COMT was successful according to the rank of the known COMT inhibitors compared to the subset of the FIMM library that was screened. I created the hitlist of 60 compounds based on the scores of these compounds, pharmacophore search and visual examination. 30 of these compounds were purchased and are currently being tested. The results of the DNMT2 run were not as reliable as the results of COMT run mentioned before, since the DNMT2 run was unable to retrieve known inhibitors better than random. The reason for that can be the quality of the model of the missing loop or the chosen controls. Furthermore only one of the ten small molecules that we used as controls is proved to be DNMT2 inhibitor, the others are DNMT1 and DNMT3 inhibitors and while the binding sites of DNMT1, DNMT2 and DNMT3 are very similar, they are, however, not completely identical.