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Nucleic Acids Research Advance Access published online on November 2, 2007
Nucleic Acids Research, doi:10.1093/nar/gkm922
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© 2007 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.

Molecular Biology

Exploring the sequence space of a DNA aptamer using microarrays

Evaldas Katilius*, Carole Flores and Neal W. Woodbury

Center for BioOptical Nanotechnology, The Biodesign Institute and the Department of Chemistry and Biochemistry, Arizona State University, Tempe, AZ 85287-5201, USA

*To whom correspondence should be addressed. Tel: +1 480 727 8790; Fax: +1 480 727 0396; Email: ekatilius{at}asu.edu

Received June 18, 2007. Revised September 18, 2007. Accepted October 10, 2007.

The relationship between sequence and binding properties of an aptamer for immunoglobulin E (IgE) was investigated using custom DNA microarrays. Single, double and some triple mutations of the aptamer sequence were created to evaluate the importance of specific base composition on aptamer binding. The majority of the positions in the aptamer sequence were found to be immutable, with changes at these positions resulting in more than a 100-fold decrease in binding affinity. Improvements in binding were observed by altering the stem region of the aptamer, suggesting that it plays a significant role in binding. Results obtained for the various mutations were used to estimate the information content and the probability of finding a functional aptamer sequence by selection from a random library. For the IgE-binding aptamer, this probability is on the order of 10–10 to 10–9. Results obtained for the double and triple mutations also show that there are no compensatory mutations within the space defined by those mutations. Apparently, at least for this particular aptamer, the functional sequence space can be represented as a rugged landscape with sharp peaks defined by highly constrained base compositions. This makes the rational optimization of aptamer sequences using step-wise mutagenesis approaches very challenging.


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