NYMC Faculty Publications

A Surface Plasmon Resonance Study of the Intermolecular Interaction Between Escherichia Coli Topoisomerase I and pBAD/Thio Supercoiled Plasmid DNA

Author Type(s)

Faculty

DOI

10.1016/j.bbrc.2014.02.015

Journal Title

Biochemical and Biophysical Research Communications

First Page

445

Last Page

450

Document Type

Article

Publication Date

3-7-2014

Department

Biochemistry and Molecular Biology

Keywords

DNA Topoisomerases, Type I, DNA, Superhelical, Escherichia coli, Magnesium, Plasmids, Surface Plasmon Resonance

Disciplines

Medicine and Health Sciences

Abstract

To date, the bacterial DNA topoisomerases are one of the major target biomolecules for the discovery of new antibacterial drugs. DNA topoisomerase regulates the topological state of DNA, which is very important for replication, transcription and recombination. The relaxation of negatively supercoiled DNA is catalyzed by bacterial DNA topoisomerase I (topoI) and this reaction requires Mg(2+). In this report, we first quantitatively studied the intermolecular interactions between Escherichia coli topoisomerase I (EctopoI) and pBAD/Thio supercoiled plasmid DNA using surface plasmon resonance (SPR) technique. The equilibrium dissociation constant (Kd) for EctopoI-pBAD/Thio interactions was determined to be about 8 nM. We then studied the effect of Mg(2+) on the catalysis of EctopoI-pBAD/Thio reaction. A slightly higher equilibrium dissociation constant (~15 nM) was obtained for Mg(2+) coordinated EctopoI (Mg(2+)EctopoI)-pBAD/Thio interactions. In addition, we observed a larger dissociation rate constant (kd) for Mg(2+)EctopoI-pBAD/Thio interactions (~0.043 s(-1)), compared to EctopoI-pBAD/Thio interactions (~0.017 s(-1)). These results suggest that enzyme turnover during plasmid DNA relaxation is enhanced due to the presence of Mg(2+) and furthers the understanding of importance of the Mg(2+) ion for bacterial topoisomerase I catalytic activity.

Link to Full Text

Share

COinS