A surface plasmon resonance study of the intermolecular interaction between Escherichia coli topoisomerase I and pBAD/Thio supercoiled plasmid DNA

Authors

Purushottam Babu Tiwari, Department of Physics, Florida International UniversityFollow
Thirunavukkarasu Annamalai, Department of Chemistry and Biochemistry, Florida International UniversityFollow
Bokun Cheng, Department of Biochemistry and Molecular Biology, New York Medical College
Gagandeep Narula, Department of Chemistry and Biochemistry, Florida International UniversityFollow
Xuewen Wang, Department of Physics, Florida International UniversityFollow
Yuk-Ching Tse-Dinh, Department of Chemistry and Biochemistry, Florida International UniversityFollow
Jin He, Department of Physics, Florida International UniversityFollow
Yesim Darici, Department of Physics, Florida International UniversityFollow

Date of this Version

3-7-2014

Document Type

Article

Rights

default

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.

DOI

10.1016/j.bbrc.2014.02.015

Identifier

24530905

Comments

Pubmed Accepted Manuscript

Published in final edited form as: Biochem Biophys Res Commun. 2014 March 7; 445(2): 445–450. doi:10.1016/j.bbrc.2014.02.015.

Recommended Citation

Tiwari PB, Annamalai T, Cheng B, Narula G, Wang X, Tse-Dinh YC, He J, Darici Y. A surface plasmon resonance study of the intermolecular interaction between Escherichia coli topoisomerase I and pBAD/Thio supercoiled plasmid DNA. Biochem Biophys Res Commun. 2014 Mar 7;445(2):445-50. doi: 10.1016/j.bbrc.2014.02.015. Epub 2014 Feb 12. PMID: 24530905; PMCID: PMC3982219.