Opening of DNA double helix at room temperature: Application of a-cyclodextrin self-aggregates† Syed S. Jaffer, a Prasun Ghosh, a Anindita Das b and Pradipta Purkayastha * a Received 10th March 2010, Accepted 30th April 2010 DOI: 10.1039/c0nr00184h Self-aggregation of a-cyclodextrin (a-CD) can induce DNA opening at room temperature (25  C) owing to the hydroxyl groups on the surface of the spherical aggregates of a-CD, which promote hydrogen bonding with the flipped-out bases in DNA duplex prohibiting them from reverting back. The process of nucleobase flipping could represent the first step in key fundamental biological processes like replication and transcription. Thus, determination of the precise mechanism of base flipping is important in understanding processes like enzyme-catalyzed DNA methylation, repair, mismatch recognition, and initiation of tran- scription and replication. 1 Such studies would also form the basis to elucidate the general principles that govern the protein–DNA inter- actions and drug development approaches targeting nucleic acids. X-Ray crystallographic studies provide the structural aspect of base flipping of cytosine 5-methyltransferase from HhaI (HhaI MTase) complexed with its DNA substrate. 1a,2 The target cytosine was found to flip completely out of the helix and into the catalytic site of the binding enzyme without seriously disturbing the rest of the DNA helix. The base flipping may be induced or facilitated by enzyme binding or alternatively, the enzyme may recognize and bind a single base transiently flipped out from the helix. 3a The rate constant of methyltransferase reaction is estimated to be 0.02 s 1 , while the measured flipping lifetime of base pair of DNA duplex is 10 ms. 3 These findings eventually indicate that an active involvement of an enzyme in accelerating base flipping appears to be unnecessary. Computational methods with full description of all chemical bonds indicate the feasibility of spontaneous base flipping in B-DNA crystal structures in the absence of an enzyme. 4 In confirmation of the theoretical findings, Spies et al. mentioned two simple models for the process by which this flipping of DNA bases occurs. 5 In one model, the enzyme catalyzes base flipping and in the other the enzyme relies on normal, spontaneous motions of the nucleic acid to provide it with a fully flipped-out target base. The spontaneously flipped-out base from double helical nucleic acids, however, can be trapped by host–guest complexation with b-cyclodextrin (b-CD). 5 Spies et al. suggest that a-CD does not have any effect on DNA melting due to insufficient space to encapsulate the nucleobases and g-CD affects the DNA very little due to weak binding. 5 According to them 50 mM b-CD lowers the melting temperature of DNA (T m ) by 1.4  C (from 62.2  C to 60.8  C) and 50 mM g-CD lowers the T m by 0.5  C. The work highlighted the encapsulation of the nucleobases inside the CD cavities and analyzed the observations. However, during the progress of research on CDs, facts like formation of self-aggregate and molecule induced nanotubular aggregates by them have been observed. 6 CDs are known to form spherical self-aggregates and the driving force for the self-assembly of cyclodextrin molecules can be considerably ascribed to hydrogen bonding. 6 Moreover, the solubility of b-CD is much less in water at room temperature (10 mM) and has a nephrotoxic effect. 7 The natural a-CD and b-CD, unlike g-CD, cannot be hydrolyzed by human salivary and pancreatic amylases. However, both a-CD and b-CD can be fermented by the intestinal microflora. a-CD and g-CD have moderately good solubility in water, but g-CD is much more expensive than a-CD and b-CD. 7a CDs have hydroxyl groups on both the rims and they form the aggregates through hydrogen bond formation. 6a Consequently, the aggregates have a good number of hydroxyl groups on the surface. So, if the nucleobases flip and the CD aggregates are close enough then they may form hydrogen bonds with the hydroxyl groups present on the surface of the aggregates (Fig. 1). Fig. 1 Nucleobase flipping out of the DNA helix (shown in green) interacts with the spherical aggregate of a-CD through hydrogen bonding. The a-CD aggregate has large number of hydroxyl groups on its surface. a Department of Chemical Sciences, Indian Institute of Science Education and Research, Kolkata, Mohanpur, 741252, WB, India. E-mail: ppurkayastha@iiserkol.ac.in b Department of Biological Sciences, Indian Institute of Science Education and Research, Kolkata, Mohanpur, 741252, WB, India † Electronic supplementary information (ESI) available: Experimental details; absorbance data of ds-DNA at higher concentration of a-CD. See DOI: 10.1039/c0nr00184h 1420 | Nanoscale, 2010, 2, 1420–1422 This journal is ª The Royal Society of Chemistry 2010 COMMUNICATION www.rsc.org/nanoscale | Nanoscale Downloaded on 13 June 2011 Published on 04 June 2010 on http://pubs.rsc.org | doi:10.1039/C0NR00184H View Online