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Nucleic Acids Research, 2000, Vol. 28, No. 8 1830-1837
© 2000 Oxford University Press

Determination of the recognition sequence of Mycobacterium smegmatis topoisomerase I on mycobacterial genomic sequences

Devanjan Sikder and Valakunja Nagaraja*

Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India

Mycobacterium smegmatis topoisomerase I has several distinctive features. The absence of the zinc finger motif found in other prokaryotic type I topoisomerases and the ability of the enzyme to recognise single-stranded and duplex DNA are unique characteristics of the enzyme. We have mapped the strong topoisomerase sites of the enzyme on genomic DNA sequences from Mycobacterium tuberculosis and M.smegmatis. The enzyme does not nick DNA in random fashion and DNA cleavage occurred at a few specific sites. Mapping of these sites revealed conservation of a pentanucleotide motif CG/TCT{downarrow}T at the cleavage site ({downarrow} represents the cleavage site). The enzyme binds and cleaves consensus oligo­nucleotides having this sequence motif. The protein exhibits a very high preference for C or a G residue at the +2 position with respect to the cleavage site. Based on earlier and the present studies we propose that the enzyme functions in vivo mainly at these specific sites to carry out topological reactions.

* To whom correspondence should be addressed. Tel: +91 80 360 0668; Fax: +91 80 360 2697; Email: vraj@mcbl.iisc.ernet.in


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