In 1999, Wan et al. [Proc. Natl. Acad. Sci. USA 96, 6014 -6019] published a pioneering paper that established the entanglement between DNA base pair motions and the transfer time of the charge carrier. The DNA assemblies contained an ethidium covalently bound via a flexible alkyl chain to the 5 hydroxyl group of the DNA backbone. Although covalently attached, the loose way in which the ethidium was linked to DNA allowed for large degrees of conformational freedom and thus raised some concern with respect to conformational inhomogeneity. In this letter, we report studies on a different set of ethidium DNA conjugates. In contrast to the ''Caltech systems,'' these conjugates contain ethidium tightly incorporated (as a base pair surrogate) into the DNA base stack, opposite to an abasic site analog. Despite the tight binding, we found that charge transfer from the photoexcited ethidium base pair surrogate across two or more base pairs is several orders of magnitude slower than in case of the DNA systems bearing the tethered ethidium. To further broaden the scope of this account, we compared (oxidative) electron hole transfer and (reductive) electron transfer using the same ethidium chromophore as a charge donor in combination with two different charge acceptors. We found that both electron and hole transfer are characterized by similar rates and distance dependencies. The results demonstrate the importance of nuclear motions and conformational flexibility and underline the presence of a base gating mechanism, which appears to be generic to electronic transfer processes through -stacked nucleic acids.conformational gating ͉ electron transfer ͉ ultrafast spectroscopy ͉ hole transfer O ver the last decade, there were many conf licting reports on the electronic conduction properties of DNA (1-4). From the multitude of studies in various laboratories, it has emerged that DNA must be understood as a dynamic medium with nuclear f luctuations, which extend over many time scales (5-11). Since the first femtosecond time-resolved experiments on ethidium (E) nucleotide complexes were reported (12), it has become clear that charge transfer through the base stack cannot be reduced to a static ''donor-bridge-acceptor'' problem. Several studies followed that highlighted the importance of conformational gating for electronic transfer processes in DNA (6 -11, 13) and peptides (14). The work presented here was motivated and inspired by the first publication on real-time DNA charge-transfer dynamics with photoexcited E in 1999 (5). The DNA systems investigated by the California Institute of Technology (Caltech) group contained E as chromophore that has been covalently attached to the 5Ј terminal hydroxyl group of one of the oligonucleotides (15). Femtosecond anisotropy measurements revealed that the f lexible alkyl chain linker warranted site-specific intercalation (within an error of 1 bp distance) without significantly restraining the orientational motions of E within the base stack (5).In most previous experiments, E has been used as...

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Fluorescent or luminescent probes that are sensitive to the local environment within DNA duplexes represent important tools for DNA hybridisation [1] and conformational changes caused by DNA±protein interactions, [1] or for the detection of physiologically important DNA base mismatches or lesions on DNA chips or microarrays.[2] As a consequence, there is a continuously increasing demand for new fluorophores that have a clear and specific range of spectral characteristics which are tunable to distinct excitation or emission wavelengths. One suitable and important way to create new emission properties is to attach chromophores covalently to natural DNA bases. Recently, we applied this modification strategy to the preparation of photoexcitable charge donors, which have been used for the investigation of DNA-mediated electron transport. [3,4] Herein, we report the properties of DNA duplexes bearing the 1-ethynylpyrene moiety (PyÀCC) covalently attached to the bases dX = dA, dC, dG, or dU. Three structural features of these PyÀCCÀdX-modified DNA duplexes are important: i) a clear steric separation of the pyrene moiety from the DNA base stack due to the rigid ethynyl group, ii) a strong electronic coupling between the pyrene and the base moiety provided by the acetylene bridge and iii) a partial stacking of the base moiety as part of the delocalised PyÀCCÀdX chromophore. Moreover, the incorporation of the PyÀCCÀdX moiety could influence only the local conformation, but should not perturb the overall B-DNA duplex conformation.The PyÀCCÀdX-modified oligonucleotides were synthesised by a semiautomated synthetic strategy with solid-phase Sonogashira-type cross-coupling conditions (Scheme 1). [4,5] It is important to note that a time-consuming synthesis of PyÀCCÀ modified phosphoramidites [6] can be avoided because this modification protocol is based on commercially available DNA building blocks. First, the oligonucleotide was synthesised by following standard protocols on a DNA synthesiser up to the position of the PyÀCCÀdX unit. At this position, either 8-bromo-2'-deoxyadenosine, 2'-deoxy-5-iodocytidine, 8-bromo-2'-deoxyguanosine, or 2'-deoxy-5-iodouridine was inserted automatically without the final deprotection of the terminal 5'-OH group. Subsequently, the CPG vials were removed from the synthesiser and a Sonogashira-coupling reagent mixture containing Pd(PPh 3 ) 4 (60 mm), 1-ethynylpyrene (120 mm) and CuI (60 mm) in DMF/Et 3 N (3.5:1.5) was added to the CPG vials under dry conditions with syringes. After a coupling time of 3 h at room temperature, the CPGs were washed with different solvents, dried and attached to the DNA synthesiser to finish the synthesis automatically. Modification of the standard procedures for deprotection and cleavage of the oligonucleotides from the solid phase, or during workup was not necessary. The PyÀCCÀdX-modified oligonucleotides were purified by semipreparative HPLC and identified by MALDI-TOF mass spectrometry. HPLC analysis of the unpurified oligonucleotides showed excellent coupling ...

Combining the fluorescence properties of phenanthridinium as an artificial DNA base together with DNA-mediated charge transfer processes allows the homogeneous detection of DNA base mismatches and abasic sites.

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