Influence of Base Stacking and Hydrogen Bonding on the Fluorescence of 2-Aminopurine and Pyrrolocytosine in Nucleic Acids

Hardman, Samantha J. O.; Thompson, Katherine C.

doi:10.1021/bi060479t

Cited by 76 publications

(89 citation statements)

References 37 publications

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“…Emission from 2-aminopurine (2AP), a structural isomer of adenine, is thought to be quenched by electron transfer with the natural bases in DNA (19,20), although this is disputed by some workers (21,22). The natural bases have comparable ionization energies and electron affinities as 2-aminopurine, and their lowest singlet excited states lie even higher in energy.…”

Section: Discussionmentioning

confidence: 99%

UV excitation of single DNA and RNA strands produces high yields of exciplex states between two stacked bases

Takaya

Su²,

Harpe³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

181

308

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Excited electronic states created by UV excitation of the diribonucleoside monophosphates ApA, ApG, ApC, ApU, and CpG were studied by the femtosecond transient-absorption technique. Bleach recovery signals recorded at 252 nm show that long-lived excited states are formed in all five dinucleosides. The lifetimes of these states exceed those measured in equimolar mixtures of the constituent mononucleotides by one to two orders of magnitude, indicating that electronic coupling between proximal nucleobases dramatically slows the relaxation of excess electronic energy. The decay rates of the long-lived states decrease with increasing energy of the charge-transfer state produced by transferring an electron from one base to another. The charge-transfer character of the long-lived states revealed by this analysis supports their assignment to excimer or exciplex states. Identical bleach recovery signals were seen for ApA, (A) 4, and poly(A) at delay times >10 ps after photoexcitation. This indicates that excited states localized on a stack of just two bases are the common trap states independent of the number of stacked nucleotides. The fraction of initial excitations that decay to long-lived exciplex states is approximately equal to the fraction of stacked bases determined by NMR measurements. This supports a model in which excitations associated with two stacked bases decay to exciplex states, whereas excitations in unstacked bases decay via ultrafast internal conversion. These results establish the importance of charge transferquenching pathways for UV-irradiated RNA and DNA in roomtemperature solution.DNA excited states ͉ DNA photodamage ͉ electronic structure ͉ femtosecond spectroscopy E xcited electronic states created in DNA by UV light have been studied since the 1960s but have received a great deal of attention recently because of advances in experiment and theory (1). These efforts are motivated by the desire to understand the photoreactions behind the genetic damage induced by UV light. There is also increasing interest in DNA and other arrays of -stacked chromophores as materials for optoelectronic applications (2-4).Striking differences have emerged between the dynamics of excited states in single bases and those in base assemblies. Excited states of single nucleobases and mononucleotides decay to the ground state primarily by ultrafast internal conversion in several hundred femtoseconds (5, 6). The additional degrees of freedom in polymeric DNA might be expected to quench singlet excited states even more rapidly, but the lifetimes actually increase dramatically. Femtosecond transient absorption measurements reveal that excited states in oligo-and polynucleotides relax in tens to hundreds of picoseconds (1, 7-10).Although there is consensus that electronic relaxation can take place orders of magnitude more slowly in DNA polymers, contradictory assignments have been proposed for the long-lived states. assigned the long-lived excited states seen in (dA) 18 , (dA) 18 ⅐(dT) 18 , and (dAdT) 9 ⅐(dAdT) 9 , to exci...

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Section: Discussionmentioning

confidence: 99%

UV excitation of single DNA and RNA strands produces high yields of exciplex states between two stacked bases

Takaya

Su²,

Harpe³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

181

308

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“…) and hydrogen bonding (1669 cm À1 ); (3) the adenine analog 2-aminopurine (abbreviated as 2AP, A* in the loop, or a* in the stem) fluoresces most strongly when base stacking is lost (Menger et al 2000;Hardman and Thompson 2006;Sarkar et al 2009). In our previous work, we looked at 4 the folding of the RNA hairpin ga*cUUCGguc (Sarkar et al 2009) after earlier studies indicated that 2AP should be an excellent reporter of folding thermodynamics and kinetics (Ballin et al 2007).…”

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confidence: 99%

Loop and stem dynamics during RNA hairpin folding and unfolding

Sarkar¹,

Nguyễn²,

Gruebele³

2010

RNA

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2-Aminopurine (2AP) is a fluorescent adenine analog that probes mainly base stacking in nucleic acids. We labeled the loop or the stem of the RNA hairpin gacUACGguc with 2AP to study folding thermodynamics and kinetics at both loci. Thermal melts and fast laser temperature jumps detected by 2AP fluorescence monitored the stability and folding/unfolding kinetics. The observed thermodynamic and kinetic traces of the stem and loop mutants, though strikingly different at a first glance, can be fitted to the same free-energy landscape. The differences between the two probe locations arise because base stacking decreases upon unfolding in the stem, whereas it increases in the loop. We conclude that 2AP is a conservative adenine substitution for mapping out the contributions of different RNA structural elements to the overall folding process. Molecular dynamics (MD) totaling 0.6 msec were performed to look at the conformations populated by the RNA at different temperatures. The combined experimental data, and MD simulations lead us to propose a minimal four-state free-energy landscape for the RNA hairpin. Analysis of this landscape shows that a sequential folding model is a good approximation for the full folding dynamics. The frayed state formed initially from the native state is a heterogeneous ensemble of structures whose stem is frayed either from the end or from the loop.

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“…Variations in AP fluorescence observed in this study are unique because they reflect the subtleties of sequence-dependent dynamic heterogeneity, not a specific conformational motif. Recent theoretical studies (37) indicate that it is the base stacking of AP, not the hydrogen bonding, that is a major factor in fluorescence quenching and that the molecular orbitals involved in fluorescence transition spread over more than one base. The quenching effect is found to be most pronounced when AP is stacked with a purine base rather than a pyrimidine base, indicating the importance of extended π-orbital conjugation (38).…”

Section: Discussionmentioning

confidence: 99%

Fluorescence Probing of Aminofluorene-Induced Conformational Heterogeneity in DNA Duplexes

Jain

Reshetnyak²,

Gao³

et al. 2008

Chem. Res. Toxicol.

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Fluorescence spectroscopy was used to study carcinogen-induced conformational heterogeneity in DNA duplexes. The fluorophore 2-aminopurine (AP) was incorporated adjacent (5′) to the lesion (G*) in eight different DNA duplexes [d(5′-CTTCTPG*NCCTC-3′):d(5′-GAGGNXTAGAAG-3′), G* = FAF adduct, P = AP, N = G, A, C, T, and X = C, A] modified by FAF [N-(2′-deoxyguanosin-8-yl)-7-fluoro-2-aminofluorene], a fluorine-tagged model DNA adduct derived from the potent carcinogen 2-aminofluorene. Steady-state measurements showed that fluorescence intensity and Stern-Volmer constants (K sv ) derived from acrylamide quenching experiments decreased for all carcinogen-modified duplexes relative to the controls, which suggests greater AP stacking in the duplex upon adduct formation. Conformation-specific stacking of AP with the neighboring adduct was evidenced by a sequence-dependent variation in fluorescence intensity, position of emission maximum, degree of emission quenching by acrylamide, and temperature-dependent spectral changes. The magnitude of stacking was in the order of FAF residue in base-displaced stacked (S) > minor groove wedged (W) > major groove B type (B). This work represents a novel utility of AP in probing adduct-induced conformational heterogeneities in DNA duplexes.

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Influence of Base Stacking and Hydrogen Bonding on the Fluorescence of 2-Aminopurine and Pyrrolocytosine in Nucleic Acids

Cited by 76 publications

References 37 publications

UV excitation of single DNA and RNA strands produces high yields of exciplex states between two stacked bases

UV excitation of single DNA and RNA strands produces high yields of exciplex states between two stacked bases

Loop and stem dynamics during RNA hairpin folding and unfolding

Fluorescence Probing of Aminofluorene-Induced Conformational Heterogeneity in DNA Duplexes

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