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DNA adopts normal B-form upon incorporation of highly fluorescent DNA base analogue tC: NMR structure and UV-Vis spectroscopy characterization

Journal article
Authors Cecilia Engman
Peter Sandin
Sadie Osborne
Tom Brown
Martin Billeter
Per Lincoln
Bengt Nordén
Bo Albinsson
Marcus Wilhelmsson
Published in Nucleic Acids Research
Volume 32
Issue 17
Pages 5087-95
ISSN 1362-4962
Publication year 2004
Published at Department of Chemistry
Pages 5087-95
Language en
Links dx.doi.org/10.1093/nar/gkh844
https://gup.ub.gu.se/file/71339
Keywords Circular Dichroism, DNA, chemistry, Fluorescent Dyes, chemistry, Nuclear Magnetic Resonance, Biomolecular, Nucleic Acid Conformation, Phenothiazines, chemistry, Spectrophotometry, Spectrophotometry, Ultraviolet, Temperature
Subject categories Biophysical chemistry, Spectroscopy, Biochemistry

Abstract

The influence of the highly fluorescent tricyclic cytosine base analogue (tC) on duplex DNA conformation is investigated. The duplex properties are characterized by absorbance and circular dichroism (CD) for all combinations of neighbouring bases to tC, and an NMR structure is determined for one tC-containing sequence. For the oligonucleotides with one tC incorporated instead of cytosine, the melting temperature is increased on average by 2.7 degrees C above that for the unmodified ones. CD spectra are practically identical for modified and unmodified sequences, indicating an unperturbed B-DNA conformation. The NMR structure determination of the self-complementary sequence 5'-CTC(tC)ACGTGGAG shows a DNA conformation consistent with B-form for the whole duplex. The root-mean-square distance for the nucleotides of the eight central base pairs between the 10 structures with lowest CYANA target functions and a mean structure is 0.45 +/- 0.17 A. The NMR data confirm correct base pairing for tC by the observation of both intrastrand and interstrand imino proton NOEs. Altogether, this suggests that tC works well as a cytosine analogue, i.e. it is situated in the base stack, forming hydrogen bonds with G in the complementary strand, without distorting the DNA backbone conformation. This first example of an artificial, highly fluorescent DNA base that does not perturb the DNA conformation could have valuable applications for the study of the structure and dynamics of nucleic acid systems.

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