© 2006 The Japanese Biochemical Society.
Regular Paper |
A Palindromic Repeat Sequence Adopts a Stable Fold Back Structure under Supercoiling
1 Centre for Biotechnology, Jawaharlal Nehru University, New Mehrauli Road, New Delhi-67, India
To whom correspondence should be addressed. E-mail: kunalrb{at}hotmail.com
A synthetic deoxyoligonucleotide containing five palindromic repeats of GGATCC self assembles to form a parallel four-stranded structure held together by G-tetrads that shows slower mobility than duplex DNA. This structure is hypersensitive to S1 nuclease and resistant to DMS modification. The same oligonucleotide when cloned in a plasmid forms a different structure under supercoiling that persists stably even in the cleaved out insert. On polyacrylamide gel electrophoresis, the cleaved out insert moves to a position midway between the duplex and parallel four-stranded forms of the oligonucleotide. Upon S1 nuclease treatment, the cleaved out insert shows a discreet band of 18 base pairs, suggesting an unfolded region in the middle. All the guanines in the cleaved out insert are sensitive to DMS modification and produce a positive peak at 285 nM in the circular dichroism spectrum, a signature of fold back tetraplex structures. We propose a fold back quadruplex structure for the insert under supercoiling with only A.T.A.T and G.C.G.C tetrads. This is the first suggestive evidence of a general tetraplex motif without G quartets as that proposed for generalized recombination.
* Present address: Department of Molecular Membrane Biology, Max Planck Institute of Biophysics, Frankfurt am main 60438, Germany. Fax: +49-69-63031002, E-mail: arun.shukla{at}mpibp-frankfurt.mpg.de