Nucleic Acids Research Advance Access originally published online on July 20, 2009
Nucleic Acids Research 2009 37(17):5589-5601; doi:10.1093/nar/gkp598
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Nucleic Acids Research, 2009, Vol. 37, No. 17 5589-5601
© 2009 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Chemistry and Synthetic Biology |
Conformationally rigid nucleoside probes help understand the role of sugar pucker and nucleobase orientation in the thrombin-binding aptamer
1Laboratory of Medicinal Chemistry, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD 21702, USA, 2DISMA, Universita’ degli Studi di Milano. Via Celoria 2, I-20133 Milano, Italy, 3Institute for Research in Biomedicine, IQAC-CSIC, CIBER-BBN Networking Centre on Bioengineering, Biomaterials and Nanomedicine, Baldiri Reixac 15, E-08028 Barcelona, 4Joint IRB-BSC program on Computational Biology. Institute for Research in Biomedicine, Baldiri Reixac 10-12, E-08028 Barcelona and Barcelona Supercomputing Center, Jordi Girona 29, 08034 Barcelona, Department of Biochemistry, University of Barcelona, Diagonal 647, 08028 Barcelona, 5Instituto de Química Física Rocasolano. CSIC, C/Serrano, 119, 28006 Madrid, Spain
*To whom correspondence should be addressed. Tel: +34 93 4039942; Fax: +34 93 2045904; Email: recgma{at}cid.csic.es
Correspondence may also be addressed to Víctor E. Marquez. Tel: +1 301 8465954; Fax: +1 301 8466033; Email: marquezv{at}mail.nih.giv
Received April 23, 2009. Revised June 29, 2009. Accepted June 30, 2009.
Modified thrombin-binding aptamers carrying 2'-deoxyguanine (dG) residues with locked North- or South-bicyclo[3.1.0]hexane pseudosugars were synthesized. Individual 2'-deoxyguanosines at positions dG5, dG10, dG14 and dG15 of the aptamer were replaced by these analogues where the North/anti and South/syn conformational states were confined. It was found that the global structure of the DNA aptamer was, for the most part, very accommodating. The substitution at positions 5, 10 and 14 with a locked South/syn-dG nucleoside produced aptamers with the same stability and global structure as the innate, unmodified one. Replacing position 15 with the same South/syn-dG nucleoside induced a strong destabilization of the aptamer, while the antipodal North/anti-dG nucleoside was less destabilizing. Remarkably, the insertion of a North/anti-dG nucleoside at position 14, where both pseudosugar conformation and glycosyl torsion angle are opposite with respect to the native structure, led to the complete disruption of the G-tetraplex structure as detected by NMR and confirmed by extensive molecular dynamics simulations. We conclude that conformationally locked bicyclo[3.1.0]hexane nucleosides appear to be excellent tools for studying the role of key conformational parameters that are critical for the formation of a stable, antiparallel G-tetrad DNA structures.
The authors wish it to be known that, in their opinion, the first four authors should be regarded as joint first authors.