alexa DNA Spiral Supercoiling and Intramolecular Topological
ISSN: 1948-5948

Journal of Microbial & Biochemical Technology
Open Access

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Research Article

DNA Spiral Supercoiling and Intramolecular Topological Interlink

Xitai Huang*, Jia Yu, Zhenfeng Zhang and Kou Cao

Department of Biochemistry and Molecular Biology, College of Life Sciences, Nankai University, Tianjin, 300071, P. R. China

*Corresponding Author:
Xitai Huang
Department of Biochemistry and Molecular Biology
College of Life Sciences, Nankai University
94 Weijin Road, Tianjin, P. R. China
Tel: +86 22 23508874
Fax: +86 22 23508874
E-mail:[email protected]

Received date: August 13, 2011; Accepted date: October 17, 2011; Published date: October 22, 2011

Citation: Huang X, Yu J, Zhang Z, Cao K (2011) DNA Spiral Supercoiling and Intramolecular Topological Interlink. J Microbial Biochem Technol S3:001. doi:10.4172/1948-5948.S3-001

Copyright: © 2011 Huang X, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

 

Abstract

Escherichia coli chromosome DNA was observed previously to consist of subunits of Archimedean spirallike supercoiling. How do cells build such a DNA structure remains unknown. In the present study, atomic force microscopy (AFM) images showed that supercoiled pBR322 DNA forms a spiral structure upon intercalation with 0.5 μg/ml ethidium bromide (EB), which used to be considered as zero superhelical density. New evidence suggested that a novel topological bond, intramolecular topological interlink (ITL), promotes the DNA spiral formation. Without intercalation, the supercoiled pBR322 DNA displays plectonemical supercoil with uneven distribution of the supercoil density. Similar observation was also made when the DNA was over intercalated by EB (20 μg/ml). The results indicated that ITL functions as a brake to block the helical double strands twisting and comparted circular DNA into different superhelical density domains. When DNA was denatured in alkaline, AFM images showed that the ITL remains constant. As the denatured pBR322 DNA was cut with restriction endonuclease PstI, the digested DNA kept intramolecular interlinks to converge into a centre with two free cut ends. Interlinked intermediates can be observed when natural pBR322 DNA was digested with HindIII and site-specific nickase Nb. Bpu10I. All evidence suggests that ITL is present in pBR322 DNA and causes DNA spiral supercoiling. It was found that the DNA topoisomers with different ITL number run into a ladder of bands in electrophoresis which is distinct from the DNA topoisomers produced by E. coli gyrase forming a smear. In a cell free system containing E. coli cell extract, we demonstrate that topoisomerase IV is required for the production of ITL DNA topoisomers from relaxed cccDNA substrate. Collectively, our data suggest that ITL represents a novel element of DNA topological structure. The DNA spiral supercoiling may be a universal structure present in the cell.

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