Studies of DNA-interacting properties of certain topological enzymes

• Main Author: Lee, Jasmine Yiqin
• Other Authors: School of Physical and Mathematical Sciences
• Format: Theses and Dissertations
• Language: English
• Published: 2013
• Subjects: DRNTU::Science::Chemistry
• Online Access: http://hdl.handle.net/10356/51880
• Institution: Nanyang Technological University

Topoisomerases are topological enzymes that are involved in the regulation of DNA supercoiling. In this thesis, two of the type IIA topoisomerases, Topoisomerase II and DNA Gyrase, will be discussed and their DNA-interacting properties will be studied. Firstly, the change in topology of Kinetoplast DNA (kDNA) catalysed by Topoisomerase II was examined. With the aid of Atomic Force Microscopy (AFM), the different forms of kDNA during the decatenation process were observed. It was found out that the optimum conditions for gel electrophoresis were set at 0.5 unit of human Topoisomerase II alpha at the reaction time of 15 minutes. Furthermore, AFM images of two minicircles and three minicircles kDNA molecules were observed. We have concluded that the decatenation process of kDNA by Topo II is a ‘step-wise’ reaction that involved one or more reaction intermediates. In the next part of the studies, the development of stable duplex oligonucleotides and its inhibitory effect on DNA gyrase were examined. Due to the increase in bacterial resistance to antibacterial agents, it is important to explore new types of antibacterial agents. One of the “dumbbell-shaped” oligonucleotides that we have designed has been shown to have inhibitory effect on DNA gyrase and has an IC50 value of 31.62 nM. In addition, it is demonstrated that the positions of the nick sites on the oligonucleotides will affect the inhibitory action on E.coli gyrase significantly. The DNA-interacting properties of the two enzymes, Topoisomerase II and DNA gyrase were explored. These studies and findings have increased our knowledge between the topological properties of DNA and these enzymes. In addition, the information provided may contribute and benefit to the future studies of these topological enzymes.