"The First Nucleic Acid Base Analogue FRET-pair - A Versatile Tool for Detailed Biochemical and Nanotechnological Structure Information"
Department of Chemical and Biological Engineering
Chalmers University of Technology
Utilization of the tricyclic cytosine family, tC, tCO, and tCnitro, is increasing in biophysical, biochemical and (bio)nanotechnological applications. The two fluorescent members of this family, tC and tCO, both have unique properties among fluorescent base analogues.[2,3] In contrast to other reported fluorescent base analogues tC and tCO have i) a high quantum yield (ff»0.2) in duplex that is virtually insensitive to neighboring base combination, ii) an emission after incorporation into DNA characterized by a single exponential decay in double stranded systems, and iii) an average luminescence brightness of the base analogues in duplex DNA being the highest reported so far and up to 50 times higher than the most commonly used fluorescent base analogue 2-aminopurine.[2,3] Furthermore, the three tricyclic base analogues all form stable base pairs with guanine and give minimal perturbations to the native structure of DNA. Importantly, we have recently utilized tCO as a donor and developed tCnitro as an acceptor and, thus, established the first nucleic acid base analogue förster resonance energy transfer (FRET)-pair. The FRET-pair successfully monitors distances covering up to more than one turn of the DNA duplex and, more importantly, we have shown that the rigid stacking of the two base analogues, and consequently excellent control of the their exact positions, results in very distinct FRET changes as the number of bases separating the donor and acceptor is varied. As a consequence of the exact positioning, this FRET-pair enables very high control of the orientation factor (k2). A set of DNA strands containing the FRET-pair at wisely chosen locations will, thus, make it possible to accurately distinguish distance- from orientation-changes using FRET. In combination with the good base analogue properties this points towards detailed studies of the inherent dynamics of nucleic acid structures. Moreover, the placement of FRET-pair chromophores inside the base stack will be a great advantage in studies where other (biomacro)molecules interact with the nucleic acid.
References:  L.M. Wilhelmsson, Q. Rev. Biophys., 43 (2010) 159.  P. Sandin, et al., Nucleic Acids Res., 33 (2005) 5019.  P. Sandin, et al., Nucleic Acids Res. 36 (2008) 157.  K. Börjesson, et al., J. Am. Chem. Soc. 131 (2009) 4288.
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