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Journal of Biomedicine and Biotechnology
Volume 2012, Article ID 735368, 11 pages
Research Article

Characterization of Flavonol Inhibition of DnaB Helicase: Real-Time Monitoring, Structural Modeling, and Proposed Mechanism

1Department of Biomedical Sciences, Chung Shan Medical University, No. 110, Section 1, Chien-Kuo N. Road, Taichung City 40201, Taiwan
2Department of Medical Research, Chung Shan Medical University Hospital, No. 110, Section 1, Chien-Kuo N. Road, Taichung City 40201, Taiwan

Received 1 February 2012; Revised 18 April 2012; Accepted 22 May 2012

Academic Editor: S. L. Mowbray

Copyright © 2012 Hsin-Hsien Lin and Cheng-Yang Huang. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


DnaB helicases are motor proteins essential for DNA replication, repair, and recombination and may be a promising target for developing new drugs for antibiotic-resistant bacteria. Previously, we established that flavonols significantly decreased the binding ability of Klebsiella pneumoniae DnaB helicase (KpDnaB) to dNTP. Here, we further investigated the effect of flavonols on the inhibition of the ssDNA binding, ATPase activity, and dsDNA-unwinding activity of KpDnaB. The ssDNA-stimulated ATPase activity of KpDnaB was decreased to 59%, 75%, 65%, and 57%, in the presence of myricetin, quercetin, kaempferol, and galangin, respectively. The ssDNA-binding activity of KpDnaB was only slightly decreased by flavonols. We used a continuous fluorescence assay, based on fluorescence resonance energy transfer (FRET), for real-time monitoring of KpDnaB helicase activity in the absence and presence of flavonols. Using this assay, the flavonol-mediated inhibition of the dsDNA-unwinding activity of KpDnaB was observed. Modeled structures of bound and unbound DNA showed flavonols binding to KpDnaB with distinct poses. In addition, these structural models indicated that L214 is a key residue in binding any flavonol. On the basis of these results, we proposed mechanisms for flavonol inhibition of DNA helicase. The resulting information may be useful in designing compounds that target K. pneumoniae and other bacterial DnaB helicases.