One-Electron Oxidation of Condensed DNA Toroids - American

Bioconjugate Chem. 2008, 19, 1235–1240

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One-Electron Oxidation of Condensed DNA Toroids: Injection-Site Dependent Charge (Radical Cation) Mobility Prolay Das and Gary B. Schuster* School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, Georgia 30332. Received January 26, 2008; Revised Manuscript Received March 18, 2008

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DNA condensates were formed by treating linear pUC19 plasmids ligated to an AQ-containing oligomer with spermidine. The condensates are toroid-shaped objects having a radius of 70 to 100 nm. Irradiation of the condensates with UV light (absorbed by the anthraquinone) causes the one-electron oxidation of the DNA and concomitant reaction at GG steps of the oligomer. Analysis of the distance dependence of the reaction at guanine in these condensates reveals a dependency on the position of the AQ. This observation is attributed to a reduction in the rate for trapping of the radical cation in the relatively dehydrated interior of the condensate.

INTRODUCTION A clear understanding of the mechanisms for oxidative damage to DNA is essential because these processes are considered to be responsible for mutations, aging, and some diseases (1). Extensive experimentation has shown that a radical cation (electron “hole”) introduced into a duplex DNA oligomer in solution by one-electron oxidation will migrate long distances by hopping before being irreversibly trapped by reaction with water or molecular oxygen at a remote nucleobase (2–5). In this process, the radical cation is self-stabilized in a distortion of the DNA and its nearby solvent and counterion environment that spreads the charge over several bases (a polaron). Thermal activation (by phonons) causes the polaron to hop from one site to a neighboring site. Trapping of the radical cation occurs at the most reactive nucleobase, which is typically a guanine, Gn-site (n ) 2, 3) or, in the absence of guanine, a thymine (6, 7). The site at which the radical cation is trapped is typically revealed by chemical or enzymatic treatment that results in strand cleavage at the damaged base (8). Most reported studies of long-distance radical cation transfer in DNA have been carried out using short oligomers in solution (9–12). For example, the UV irradiation of an anthraquinone group (AQ) that is covalently linked to a 5′-terminus of a duplex DNA oligomer results in oxidative damage to remote bases. We recently reported the results from a set of experiments in which DNA from a linearized plasmid was ligated to an AQcontaining oligomer, formed into a condensate, and then irradiated (13). Condensates are nanometer-scale particles produced from extended DNA chains in the presence of multivalent cations such as spermine or spermidine (14–19). They are sometimes used as models for studying gene packing in viruses, bacteria, or eukaryotic cells. Our previous investigation showed that the charge migration effectiveness decreased in the condensate compared with that in a similar short oligomer in solution. This observation was attributed to a reduction in the formation of charge-transfer effective states in the condensate (13). In that study, by design, the AQ group was at a terminal position of the duplexes that comprise the condensate. Clearly, those are unique positions and the reactions of radical cations near those sites may not represent the properties of the majority of the nucleobases in the condensate. In the current study, we prepared condensates from an oligomer that is doubly ligated to two linearized pUC19 plasmids. This construction causes the AQ group to be in an interior position of the condensate. The

pattern of oxidative damage in this system reveals a complex competition between charge hopping and charge trapping within the condensate.

EXPERIMENTAL PROCEDURES Materials and Methods. The pUC19 plasmid, T4 DNA ligase, HindIII, EcoRI, PaeR7 I restriction endonucleases, and T4 polynucleotide kinase were purchased from New England Biolabs. [γ-32P]ATP and nonradioactive ATP were purchased from Amersham Biosciences. Gel extraction kits were purchased from Qiagen. Gelase agarose gel-digesting preparation was obtained from Epicenter Biotechnologies, and spermidine hydrochloride was obtained from Aldrich. DNA oligomers and the uridine AQ were synthesized by standard procedures on an Expedite 8909 DNA synthesizer (20). The phosphorylation of the 5′-end of DNA2 was achieved in the DNA synthesizer with chemical phosphorylation reagent II, obtained from Glen Research (Sterling, VA). The oligonucleotides were purified by reverse-phase HPLC on a Hitachi (Tokyo) preparative HPLC system using a Dynamax C18 column. Purified oligomers were desalted with Sep-Pak columns and characterized by mass spectroscopy. UV melting and cooling curves were recorded on a Cary 1E spectrophotometer (Varian) equipped with a multicell block, temperature controller, and sample transport accessory. Buffer-exchanging spin columns, enzyme removal, and other spin columns are obtained from Millipore Corporation (Bedford, MA). Restriction Digestion and Alkaline Phosphatase Treatment of Plasmid. A 2 µg sample of pUC19 plasmid was treated with 5 µL of HindIII restriction endonuclease in a microcentrifuge tube (total volume of 50 µL) containing 1 × HindIII buffer and incubated overnight at 37 °C. To this solution, 0.5 µL sample of shrimp alkaline phosphatase (SAP) was added at the same restriction buffer concentration. The reaction mixture was incubated at 37 °C for an additional 30 min, followed by heating at 65 °C for 15 min. The SAP-treated restriction digest was passed through a Micropure-EZ centrifugal filter device to eliminate the restriction enzyme and SAP. The restriction buffer was replaced with nanopure water by means of buffer-exchange spin columns. The linearized plasmid samples were precipitated with ethanol, washed twice with 80% ethanol, and air-dried. The SAP-treated linearized plasmid was resuspended in water, and its concentration was measured by UV-vis spectroscopy. The same procedure was carried out for restriction digestion of

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Figure 1. A schematic representation of the experimental design for the DNA constructs used in this work. pUC19 is cut with Hind III enzyme and ligated to the HindIII compatible ends of DNA1/ DNA2, to form SLP. Further ligation of SLP with EcoRI linearized pUC19 give DLP. Treatment of SLP and DLP with spermidine leads to toroidshaped condensate formation, DNA1/DNA2 is a double-stranded DNA oligomer that has two cohesive ends; one of them is complementary to Hind III cut linearized pUC19 (shown as dark yellow) and the other complementary to EcoRI cut linearized pUC19. The oligomer duplex also has an AQ group linked covalently to a uridine (AQ, structure shown), a 5′-32P radiolabel, and a palindromic sequence which is cut by Pae R7 I (shown as canary yellow).

pUC19 with EcoRI enzyme and SAP treatment. In all cases, analytical, nondenaturing, low-melting agarose gels were run to identify and compare the linearized plasmids from uncut relaxed and supercoiled plasmid and also to confirm inhibition of self-ligation by SAP. The single- (SLP) and double- (DLP) ligated plasmid condensates were treated with PaeR7 I restriction endonuclease after irradiation. The restriction digestions were carried out in the presence of NEB buffer 4 and were supplemented by 5 µg of bovine serum albumin and incubated at 37 °C for 1 h. The reaction mixtures were used without further purification for subsequent piperidine treatment. Preparation of Radiolabeled DNA. The DNA1 oligomer (Figure 1) was radiolabeled with [γ-32P] ATP and T4-polynucleotide kinase enzyme (PNK) and isolated by standard methods on a 20% denaturing polyacrylamide gel. The DNA was precipitated by addition of cold ethanol, vortexed, placed on dry ice, and then centrifuged. The DNA sample was washed twice with 80% ethanol and then air-dried. The air-dried DNA pellets were redissolved in water and passed through bufferexchange spin columns to eliminate ammonium, magnesium, and acetate ions from the gel elution buffer. Ligation of Plasmids with Radiolabeled Oligonucleotides. Equimolar concentrations (3 µM) of radiolabeled DNA1 and complementary sequence DNA2 were mixed in 10 mM sodium phosphate buffer at pH 7.0, annealed by heating for 10 min at 90 °C, and then cooled very slowly to room temperature. The oligomer was ligated to the Hind III linearized pUC19 plasmid (10:1 molar ratio) with T4 DNA ligase, 10 mM ATP in 1 × T4 ligase buffer at 16 °C overnight in a Neslab RTE-7 incubator (Newington, NH). The success of the ligation reaction was assessed by autoradiography on an analytical, nondenaturing low-melting agarose gel following standard procedures (21). The DNA in the reaction mixture was washed with ethanol and mixed with nondenaturing dye containing bromophenol blue and xylene cyanol in water and glycerol (4:1). The ligated

Das and Schuster

plasmid DNA band was isolated from a nondenaturing 1% preparative agarose gel in 1 × TAE buffer (Tris-acetate-EDTA) containing ethidium bromide for visualization. The gel pieces were collected in centrifuge tubes and heated at 65 °C for 10 min, followed by addition of 1 µL of 50 × reaction buffer of gelase agarose gel-digesting enzyme, per 50 µL of molten agarose. The solutions were incubated overnight after the addition of one unit of gelase enzyme per 200 mg of molten agarose. The gelase digestions were washed with ethanol and then passed through Qiagen nucleotide removal spin columns for elimination of the last traces of agarose. The gel-extracted SLP was subjected to a second phase ligation by the addition of equimolar amounts of linearized pUC19 plasmid, cut with EcoRI restriction enzyme, in the presence of T4 DNA ligase and 10 mM ATP in 1 × T4-ligase buffer at 20 °C. Preparative agarose gel and gel extraction procedures, as above, were performed to obtain purified doubleligated plasmid (DLP). Dynamic Light Scattering Study. A solution of doubly ligated plasmid containing the sandwiched oligomer duplexDNA1/DNA2 (25 µg) was added to an equal volume of spermidine solutions (50-600 µM at increments of 50 uM). All experiments were performed in 10 mM sodium phosphate buffer (pH 7.2). All solutions were filtered through Amicon UltraFree MC centrifugal filters (diameter, 0.22 µm; Millipore) before introducing them into the light scattering instrument. After 5 min of gentle agitation, the samples were analyzed by dynamic light scattering using a Dynapro MS X dynamic lightscattering instrument (Proterion, Piscataway, NJ). Decondensation experiments were performed by addition of NaCl (final concentration 2 M) to the solutions containing the DNA and spermidine at appropriate concentrations. Transmission Electron Microscope Studies. DNA condensates were prepared according to the standard procedure (22); doubly ligated plasmids were deposited on a carbon-coated copper grid (Ted Pella, Redding, CA) and allowed to settle for 10 min. The grids were stained by the direct addition of 5 µL of a 2% uranyl acetate solution, rinsed in 95% ethanol, and then air-dried. Images of the DNA condensates at room temperature were collected on film using a JEOL-100C transmission electron microscope at 100 000× magnification. UV Irradiation and Strand-Cleavage Analysis. DNA solutions of the radiolabeled oligomer duplex, SLP and DLP were irradiated in microcentrifuge tubes (20 µL, 10 mM sodium phosphate buffer) containing varying concentrations of spermidine at ca. 30 °C in a Rayonet Photoreactor (Southern New England Ultraviolet, Barnsford, CT) equipped with 16 × 350 nm lamps. After irradiation, the samples were washed with ethanol, dried, and treated at 90 °C with 1 M piperidine for 30 min. The piperidine was evaporated, and the samples were dried and suspended in denaturing loading buffer and analyzed by electrophoresis on a 20% 19:1 acrylamide/bis-acrylamide containing 7 M urea Tris-borate-EDTA buffer. Cleavage sites were visualized by autoradiography and quantified by phosphorimagery.

RESULTS Preparation of Internally Linked AQ-DNA. We constructed a linear DNA oligomer (DLP; see Figure 1) that is composed of an AQ-containing duplex oligomer (DNA1/DNA2) linked at both ends to linearized pUC19 plasmids. The duplex is formed from strands DNA1 and DNA2 and it has two cohesive ends. The 5′-end of DNA1 has a 5′-AATT-3′ overhang that is compatible with the EcoR1 restriction-cut ends of pUC19. This 5′-end also contains a [32P]-radiolabeled phosphate group that will permit visualization of the experimental results by autoradiography. The 3′-end of DNA1 forms a four-base

One-Electron Oxidation of Condensed DNA Toroids

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Figure 3. Dynamic light scattering experiment: Plot of the intensity of scattered light against the concentration of spermidine in DLP.

Figure 2. (A) Nondenaturing low melting agarose gel showing the Rf of DNA1/DNA2, SLP, and DLP. Lane 1: The reaction mixture from ligation of DNA1/DNA2 with Hind III cut linearized plasmid. Lane 2: The reaction mixture from ligation of gel-extracted SLP and EcoRI cut linearized pUC19. Lane 3: 1 Kb DNA ladder. Lane 4: The gel extracted DLP. Lane 5: The gel extracted SLP. Lane 6: pUC19 control, cut with Hind III restriction endonuclease. Lane 7: circular pUC19 in supercoiled and relaxed form. (B) Autoradiography of low-melting agarose gel showing DNA1/DNA2, DLP, and SLP. Lane 1: The reaction mixture from ligation of DNA1/DNA2 with Hind III cut linearized plasmid. Lane 2: The reaction mixture from ligation of SLP with EcoRI cut linearized plasmid. Lane 3: Isolated and purified SLP. Lane 4: Isolated and purified DLP.

recessed cohesive end in the DNA1/DNA2 duplex that is complementary to the overhang of a HindIII cut pUC19 plasmid. DNA1 also has an AQ group linked covalently to a uridine in a central location. In addition, the DNA1 strand contains six GG steps (three on each side of the AQ) that will serve as indicator sites for the reactions of the radical cation. The DNA1/DNA2 duplex was prepared by following standard techniques that have been reported previously (12). The melting temperature (Tm) for this AQ-containing duplex is 68 °C, which is 3.5 °C higher than that of the corresponding duplex where the uridine-linked AQ has been replaced by a thymine. This indicates that the AQ group stabilizes the duplex oligomer, probably by intercalation (20). DLP was prepared in two steps from DNA1/DNA2. In the first step, ligation of DNA1/DNA2 with HindIII linearized pUC19 gives SLP (see Figure 1). The HindIII cuts pUC19 at a unique position (base number 447) leaving two overhanging 5′-AGCT-3′ cohesive ends. This 2680 base pair linearized plasmid was treated with shrimp alkaline phosphatase (SAP) before reaction with DNA1/DNA2 to inhibit self-ligation. The T4 DNA ligase catalyzed reaction of DNA1/DNA2 with the HindIII cut plasmid gives the SLP construct in ca. 50% yield having an AQ group close to its terminus. SLP was purified by preparative agarose gel electrophoresis to remove unligated and self-ligated DNA1/DNA2 (see Figure 2). In the second step of formation of DLP, pUC19 linearized by reaction with EcoR1, which cuts the plasmid at the unique 5′-AATT-3′ sequence site (base number 396), was ligated (after treatment with SAP) to SLP to give DLP, which was further purified by gel electrophoresis. Thus formed, the DLP construct is a linear DNA polymer containing 5407 base pairs that has the DNA1/DNA2 AQ-containing oligomer in the central position between the two linearized pUC19 plasmids. Finally, it should be noted that the HindIII compatible end of DNA1/DNA2 contains a 5′-CTCGAG-3′/3′-GAGCTC-5′ restriction site for PaeR7 endonuclease. This sequence does not occur in pUC19. This fact allows for the selective disassembly of the SLP and DLP constructs for analytical purposes after their irradiation.

Figure 4. Transmission electron microscopic image of DNA condensate formed from DLP at pH ) 7 with 100 µM spermidine in 1 × Tris-EDTA buffer. Scale indicates 100 nm. Images are taken at 100 000 magnification in a JEOL 100C TEM instrument.

Formation of DNA Condensates from DLP with Spermidine (DLP-Condensate). It is well-known that treatment of DNA with polycations such as spermidine can lead to the formation of condensate structures (14, 15). We converted DLP to condensates in this way and characterized those structures by dynamic light scattering and by transmission electron microscopy. Dynamic light scattering experiments were conducted to assess the effect of spermidine on SLP and DLP. Solutions of purified SLP and DLP oligomers were prepared with various concentrations of spermidine, and their hydrodynamic radii were measured by light scattering. In the absence of spermidine, solutions of SLP and DLP do not show meaningful scattering. The data for DLP are shown in Figure 3; the behavior of SLP (not shown) is essentially identical to that of DLP. With the addition of spermidine, the light scattering intensity of DLP solutions increases, indicating that particles are being formed and their hydrodynamic radii increase with the spermidine concentration. At spermidine concentrations between 25 and 50 µM, the light scattering intensity is low and derived particle radii ranges 40-50 nm. With an increase in spermidine concentration to the range between 75 and 125 µM, the light scattering intensity increases and then reaches a plateau, which indicates the formation of stable particles having calculated radii of 70 to 90 nm. We refer to these particles as DLP-condensates. At higher spermidine concentration, the light scattering experiment indicates the formation of larger aggregates that eventually precipitate from solution. As expected (17), the light scattering experiments show that condensate formation is reversed by the addition of NaCl (2 M) to the solution of DLP and spermidine. The formation of condensates by DLP with the spermidine concentration at 100 µM was confirmed by transmission electron microscopy (Figure 4), which reveals stable, toroid-like structures with radii of 80-100 nm.



Das and Schuster

results in an increased distance dependence for reaction at GG steps; that is, GG damage decrease more sharply with distance from the AQ for SLP-condensate compared with DNA1/DNA2 and SLP. This finding is consistent with previous experiments in which the AQ group is at the terminal position of the DNA that forms the condensate (13). Comparison of the relative reactivity of radical cations at the GG steps of DLP and DLP-condensate provides a remarkable finding. Unlike the case for SLP, condensate formation has little effect on the distance dependence of reaction for DLP (see Figure 5). A close look at the GG damage pattern for DLPcondensate (in the presence of 100 µM of spermidine) reveals that reactions at the guanines are essentially identical to those observed for linear DLP (no spermidine) and for DNA1/DNA2 with or without spermidine present. Evidently, the position of the AQ in the condensate has a powerful affect on the distance dependence of radical cation migration efficiency. When the AQ is at or near the end of the DNA strands that comprise the condensate, remote reaction of GG steps is inhibited, but when the AQ at the middle of the DNA stand, condensate formation has no measurable effect on the efficiency of remote reaction. Figure 5. Normalized damage ratios for DNA1/DNA2, DLP, and SLP irradiated in the presence or absence of spermidine, as indicated. Each bar corresponds to a GG step. The AQ is between GG3 and GG4. Damage ratios for DNA1/DNA2 are after 45 min of irradiation and those for DLP and SLP are after 2 h of irradiation.

UV Irradiation of UAQ-DNA1/DNA2, SLP, and DLP. It is known that UV irradiation of an AQ that is covalently linked to duplex DNA at an internal position introduces a nucleobase radical cation and results in reaction far removed from the anthraquinone (20). Those reactions are routinely revealed as strand cleavage at remote GG sites by treatment of the irradiated sample with piperidine (23). We carried out experiments on DNA1/DNA2, SLP, SLP-condensate, DLP, and DLP-condensate to assess the effect of condensate formation on the migration and reaction of the radical cations introduced by UV irradiation of the covalently linked AQ group. Solutions of DNA1/DNA2, DLP, and SLP, with and without spermidine, were irradiated with UV light to low conversion (single hit conditions) and then analyzed to assess the amount of reaction that occurred at each of the GG steps of the DNA1/ DNA2 oligomer. Analysis by high-resolution PAGE requires the deconstruction of the ligated plasmid. This was accomplished by its treatment with endonuclease PaeR7 I and Sac I, which excises the oligomer portion from the plasmid-oligomer-plasmid system. These results are shown in the form of a histogram in Figure 5. Autoradiograms from the PAGE analyses of irradiated samples of SLP and DLP after deconstruction and treatment with piperidine are shown in Supporting Information (Figures S1 and S2). These experiments show that UV irradiation and piperidine treatment of DNA1/DNA2, DLP, and SLP result in strand cleavage at GG steps on both the 3′ and 5′ sides of the AQ in each of the constructs both in the linear form and as the condensate. As has been previously observed, the efficiency of reaction to the 5′ side of the AQ is greater than to its 3′ side (20). This is true for DNA1/DNA2, as well as SLP and DLP. As expected, the efficiency of strand cleavage generally falls off the farther the GG step is located from the AQ. As a control experiment, solutions of DNA1/DNA2 containing spermidine were irradiated and analyzed. Piperidine treatment and PAGE analysis of the irradiated samples show (see Figure 5) that the presence of the spermidine (100 µM) does not affect the relative reactivity of the GG steps of the oligomer. In contrast, comparison of the results from irradiation of SLP without added spermidine to those containing 100 µM of spermidine reveals a strand cleavage pattern showing that condensate formation

DISCUSSION The one-electron oxidation of DNA generates a radical cation that migrates by a thermally activated hopping process requiring local structural distortion of DNA (24). These local structural distortions include the redistribution of surrounding water molecules and counterions, which are responsible for stabilizing the self-trapped radical cation (25). If the rate for hopping of the radical cation (khop) is greater than for its irreversible trapping (ktrap) at guanine-rich regions of the DNA, then the radical cation will be distributed approximately equally (thermodynamic control) to all equivalent sites. However, if the rate of trapping of the radical cation is greater than that for hopping, then the probability of reaction at remote sites typically falls off exponentially with distance (kinetic control) (5). In an earlier report (13), we showed that when the AQ is located at a terminal position the efficiency of charge hopping decreases significantly when linear DNA is converted to a toroidal condensate by the addition of spermidine. In the present case, we find a similar result for SLP-condensate, which has the AQ group near the terminus of the DNA chain. However, no such effect is observed for DLP-condensate where the AQ group is in the center of a long DNA chain. Clearly, the efficiency of radical cation hopping is affected by an environmental change associated with condensate formation that is different near the ends of the chain than it is at the middle. It is not possible to reach a unique explanation for this observation from these experiments because their outcome is determined by the ratio of the rates of hopping and trapping (5). That is, the observed effect might be a result of an increase in the magnitude of the rate of hopping near the center of the condensate, or it could be a result of the trapping rate becoming smaller (or some combination of these two effects). Nevertheless, valuable insight into the nature of radical cation migration and trapping in complex DNA structures results from these findings. In SLP, where the AQ group is 22 base pairs from a terminus, the GG damage falls off normally with distance. Evidently, positioning the AQ group at an internal position near a terminus does not change the behavior compared with the case when the AQ is at the terminus, as it is in the previously reported example (13). Thus, the factors responsible for differentiation of the behavior of SLP and DLP-condensates cannot be directly tied to the specific nature of the chemical linkage of the AQ to the DNA strand. It has been reported that the surface-to-surface spacing between adjacent DNA helices in condensed structures is ca.


One-Electron Oxidation of Condensed DNA Toroids

1-2 water molecules in diameter (26). This indicates that the DNA molecules that comprise condensates are considerably dehydrated compared with DNA oligomers in solution. Thus, there is a relative absence of water in the interior of the DLPcondensate. Water and to a lesser extent O2 are the reagents responsible for the trapping of the radical cation that leads eventually to strand cleavage. A lower concentration of these trapping reagents in the interior of the condensate would lead to a decrease in the rate of trapping of the migrating radical cation. As a result of this factor, the radical cations formed in dehydrated regions of the condensate would migrate with reduced distance dependence compared to those formed in wellhydrated regions. This explanation accounts for the findings with the DLP-condensate if it is assumed that the interior regions of the condensate are less hydrated, which seems reasonable. Interestingly, it has been suggested that the amazing resistance to radiation damage exhibited by the bacterium Deinococcus radiodurans is related to toroid-like structures found in its DNA that inhibit diffusion (27). Alternatively, the reduced distance dependence observed for DLP-condensate might be caused by an increase in the rate of hopping, although this seems less likely. Multivalent cations such as spermidine not only cause condensation but also modify the local structure of the DNA (28). If these local structural changes are different in the interior of the condensate than near the terminus of a DNA chain, and if these changes accelerate the rate of radical cation hopping, then these factors could account for the difference we observe between SLP and DLP condensates. Although it seems less likely, this explanation can not be ruled out with the available data.

CONCLUSION DNA condensates were formed from constructs created by ligating an AQ-containing oligomer to one or two linearized pUC19 plasmids. In the first case (SLP), the oligomer is near a terminal position of the DNA chain. In the second case (DLP), the oligomer is in the central position between two linearized plasmids. Both constructs are converted to condensates when they are treated with spermidine. The condensates were characterized by dynamic light scattering and TEM and found to be toroid-shaped objects having a radius of 70 to 100 nm. Irradiation of the condensates with UV light (absorbed by the AQ) causes the one-electron oxidation of the DNA resulting in the conversion of a nucleobase to its radical cation. The radical cation migrates along the DNA chain and is trapped at GG steps where it reacts irreversibly with H2O or O2. The site of reaction is revealed (after enzymatic deconstruction of the condensate) by autoradiography. Analysis of the distance dependence of radical cation migration in these condensates reveals a dependency on the position of the AQ. The radical cation migrates more efficiently in DLP-condensate than in SLP. This observation is attributed to a reduction in the rate for trapping of the radical cation in the relatively dehydrated interior of the DLP condensate compared with SLP. Alternatively, this effect could be due to a unique local structural change in the interior of the DLP condensate that increases the rate of radical cation hopping. These findings illuminate the challenge of extrapolating results obtained for linear DNA oligomers in solution to more complex DNA structures.

ACKNOWLEDGMENT We thank Prof. Andrew Lyon (School of Chemistry and Biochemistry, Georgia Institute of Technology) for assistance with dynamic light scattering experiments; Dr. Sriram Kanvah for synthesizing the DNA oligomer strands. This work was supported by a grant from National Science Foundation and by the Vassar Woolley Foundation.

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Supporting Information Available: Autoradiogram showing results from the irradiation of DNA1/DNA2, DLP and SLP and their condensates are shown in Figures S1 and S2.This material is available free of charge via the Internet at http://pubs.acs.org.

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