Novel Bifunctional Viologen-Linked Pyrene Conjugates: Synthesis and Study of Their Interactions with Nucleosides and DNA Mahesh Hariharan, Joshy Joseph, and Danaboyina Ramaiah* Photosciences and Photonics, Chemical Sciences and Technology DiVision, Regional Research Laboratory, Council of Scientific and Industrial Research, TriVandrum 695 019, India ReceiVed: May 19, 2006; In Final Form: September 27, 2006 With the objective of developing efficient DNA oxidizing agents, a new series of viologen-linked pyrene conjugates with the general formula PYLnV 2+ , having a different number of methylene spacer units (Ln) was synthesized, and their interactions with nucleosides and DNA have been investigated through photophysical and biophysical techniques. The viologen-linked pyrene derivatives PYL1V 2+ (n ) 1), PYL7V 2+ (n ) 7), and PYL12V 2+ (n ) 12) exhibited characteristic fluorescence emission of the pyrene chromophore centered around 380 nm but with significantly reduced yields when compared to those of the model compound PYL1Et 3 + . The fluorescence quenching observed in these systems is explained through an electron-transfer mechanism based on a calculated favorable change in free energy (ΔG ET )-1.59 eV), and the redox species characterized through laser flash photolysis studies. Intramolecular electron-transfer rate constants (k ET ) were calculated from the observed fluorescence yields, and the singlet lifetimes of the model compound and are found to decrease with increasing spacer length. The DNA binding studies of these systems through photophysical, chiroptical, and viscometric techniques demonstrated that these systems effectively undergo DNA intercalation with association constants (K DNA ) in the range of 1.1-2.6 × 10 4 M -1 and exhibit 2:1 sequence selectivity for poly(dG)‚poly(dC) over poly(dA)‚poly(dT). Photoactivation of these systems initiates electron transfer from the singlet excited state of the pyrene chromophore to the viologen moiety followed by an electron transfer from DNA to the oxidized pyrene. This results in the formation of stable charge-separated species such as radical cations of both DNA and reduced viologen as characterized by laser flash photolysis studies and subsequently the oxidized DNA modifications. These novel systems are soluble in buffer media, stable under irradiation conditions, and oxidize DNA efficiently and selectively through a cosensitization mechanism and hence can be useful as photoactivated DNA cleaving agents. 1. Introduction Design of functional molecules that bind selectively to DNA and are capable of modifying duplex or single-stranded DNA is an active area of research that has important biochemical and medicinal applications. 1 Several molecules, which induce DNA modifications by various mechanisms, have been reported in the literature. 2 Among these, the photoactivated DNA oxidizing agents have been found to possess significant practical advan- tages over the reagents that cleave DNA under thermal condi- tions. 3 An interesting aspect of these agents is that they allow the reaction to be controlled spatially and temporally by combining all of the components of the reaction mixture before the irradiation. Excitation of the reaction mixture with an appropriate light source initiates the reaction, which continues until the light is shut off. 4,5 The ability to control light in both spatial and temporal senses would be advantageous for various biological applications. By absorption of light, these reagents are known to modify DNA through different mechanisms, including the electron- transfer reaction, generation of diffusible and nondiffusible reactive intermediates, and hydrogen atom abstraction. 3-6 In the latter two processes, selectivity of the DNA cleavage is rather difficult to attain, as these reactions are generally nonspecific, while the former mechanism is shown to have base selectivity. A large number of simple organic as well as inorganic sensitizers have been reported that oxidize DNA through a photoinduced electron-transfer mechanism. 7,8 However, most of these sensitiz- ers were found to be less efficient due to the existence of efficient electron back-transfer between the resultant oxidized DNA and the reduced sensitizer. To overcome the drawbacks of the electron back-transfer process associated with such systems, a few examples based on a cosensitization mechanism have been developed. 9,10 These systems consist of a sensitizer, which is also an intercalator, that transfers an electron upon excitation to a cosensitizer (electron acceptor), which is bound on the surface of DNA. The photosensitization involving the cosensitizer that bound far away from the sensitizer is expected to inhibit the electron back-transfer and thereby increase the DNA modifications. However, in reality, only a marginal improvement in DNA oxidation was observed using these systems owing to the complications with respect to the concentration, distance, and DNA binding affinities of the sensitizer and cosensitizer. Therefore, molecules that exhibit considerable DNA binding affinity and specificity in cleavage are yet to be achieved. Recently, we have reported DNA binding and cleaving efficiencies of a few bifunctional conjugates consisting of acridine as the sensitizer and the viologen moiety as the cosensitizer. 11 These molecules exhibited a high affinity for * Author to whom correspondence should be addressed. Phone: (+91) 471 2515362. Fax: (+91) 471 2490186 or 2491712. E-mail: rama@csrrltrd.ren.nic.in or d_ramaiah@rediffmail.com. 24678 J. Phys. Chem. B 2006, 110, 24678-24686 10.1021/jp063079o CCC: $33.50 © 2006 American Chemical Society Published on Web 11/09/2006