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J. Phys. Chem. B 2004, 108, 15392-15397
NMR Studies of Molecular Mobility in a DNA-Amphiphile Complex Cecı´lia Leal,* Daniel Topgaard,*,§ Rachel W. Martin,§ and Håkan Wennerstro1 m Physical Chemistry1, Lund UniVersity, POB 124, 221 00 Lund, Sweden and Materials Sciences DiVision, Ernest Orlando Lawrence Berkeley National Laboratory and Department of Chemistry, UniVersity of California, Berkeley, California 94720 ReceiVed: May 6, 2004; In Final Form: June 21, 2004
The molecular mobility in a hexagonal DNA-cationic surfactant complex is studied using 1H and 13C nuclear magnetic resonance spectroscopy. The charge-compensated complex can swell in water up to a content of approximately seven water molecules per charge. The NMR measurements show that in the dry state the alkyl chains of the surfactant have the properties of a disordered solid with internal motions of sufficient amplitude to substantially narrow the 1H resonance line from the rigid lattice limit. As water is introduced, there is an increase in molecular motion resulting in further narrowing of the signal. In the fully swollen system, the signal is narrower than that observed for a normal hexagonal liquid crystalline phase with the same surfactant. This shows that the alkyl chains are packed with a degree of disorder that is higher than in the corresponding liquid crystalline surfactant system, reflecting the aggregate deformations induced by the requirement of charge matching with DNA. Furthermore, the translational diffusional motion of the surfactant molecule is slower than D < 10-13 m2/s, while for the water molecules we observe D going from 1 × 10-11m2/ s at 5 water molecules per base pair to 2 × 10-10 m2/s at the swelling limit of 27 waters per base pair. The DNA remains solid throughout the hydration range. By combining the NMR observations with the thermodynamic characterization of the system by Leal et al.1 we arrive at a detailed description of the molecular organization in the complex between DNA and the single chain cationic surfactant hexadecyltrimethylammonium, CTA.
Introduction The DNA double helix is highly negatively charged and it interacts strongly with oppositely charged species. In the intracellular environment, the DNA is associated with cationic histone proteins resulting in the formation of compact chromatin aggregates. Clearly, attractive electrostatic forces are important in promoting this compact structure. In vitro, an analogous compaction of the DNA can occur on association with cationic amphiphiles.2-4 Interestingly, studies of such systems show that hydrophobic forces are also involved and can be decisive in determining the physicochemical properties and structure of such complexes.1,4,5 Currently there is a considerable interest in DNA-amphiphile complexes due to the possibility of applying such nonviral assemblies as vehicles for delivery of foreign DNA into cells.6,7 To control the delivery process, it is essential to understand how the amphiphile self-assembles in the presence of DNA and its molecular properties in the complexes. These contain substantial amounts of water, and the hydration of the charged DNA and lipid groups is an important factor for determining the molecular organization in the complexes.1 Depending on the molecular character of the amphiphile, both lamellar and hexagonal structures have been found for chargecompensated DNA/amphiphile aggregates.1-3 Lamellar DNAlipid systems have been extensively studied with respect to both molecular organization and thermodynamic properties.8-11 It is found that the lipid properties are qualitatively similar to those of pure lipid lamellar systems, but the presence of the DNA results in substantial quantitative changes. * Corresponding authors. E-mails:
[email protected] and
[email protected]. § University of California, Berkeley.
Single-chain amphiphiles, which by themselves would typically form micelles and normal hexagonal structures, can be used more readily than the lamellar-forming lipids to produce complex particles containing only a single compacted DNA molecule. The packing in such particles is expected to resemble the structure of a normal hexagonal phase, and for a system formed by precipitating such particles one observes a smallangle X-ray pattern typical for a 2-D hexagonal structure.5 Recently, we studied thermodynamic hydration properties of such precipitated complexes. We concluded that as the water content decreases there is a gradual change in the properties of the hydrocarbon chains and that in the dry state the surfactant aggregate is highly deformed to accomplish local charge neutralization with respect to the anionic phosphate groups on the DNA.1 In the present paper, we report 1H and 13C NMR studies of the same system used in the calorimetric investigations. NMR is a useful, noninvasive tool to study the molecular properties in self-assembled systems in general.12 We use the line shape and width of the 1H signal from the amphiphile and the DNA, detected either directly or after cross polarization (CP)13 to 13C during magic angle spinning (MAS),14 to monitor changes in molecular mobility as the water content is varied. Additionally, measurements of the amphiphile and water selfdiffusion provide crucial information about the translational motion in the complex. Materials and Methods Materials. We used sodium salt DNA: Calf thymus (fibrous type 1 “highly polymerized”). It was obtained from Sigma and used as received. The concentration of DNA was determined by a spectroscopic method using the molar extinction coefficient
10.1021/jp0480495 CCC: $27.50 © 2004 American Chemical Society Published on Web 09/04/2004
Molecular Mobility in a DNA-Amphiphile Complex 260 ) 6600 L mol-1 cm-1 at 260 nm.15 The A260/A280 ratio of DNA solutions was found to be 1.9, suggesting that the DNA was free of proteins.15 Hexadecyltrimethylammonium bromide, CTAB, (Merck, p.a. quality) was used as received. The water used was from a Milli-Q filtration system (Millipore). D2O was obtained from Dr. Glaser AG, Basel. Sample Preparation. DNA-surfactant stoichiometric aggregates were prepared by mixing equimolar solutions of calf thymus DNA and CTAB under stirring. The number of cationic surfactants equals the number of phosphate groups on the DNA. The precipitate formed was equilibrated in solution for 48 h. It was then separated from the aqueous phase by filtration at reduced pressure, and washed extensively with Millipore water. The macromolecular complex salt (DNA-CTA) was dried for 3 days and equilibrated at different relative humidities in desiccators with saturated salt solutions at 25 °C. When wet, the DNA-CTA precipitate is like a paste, when dry it shows a granular morphology. For the self-diffusion and wide-line 1H experiments, the samples were placed in a 5-mm tube and well squeezed into the bottom of the tube. For the MAS experiments, dry sample was put into 3.2-mm zirconia rotors (Varian Inc.) while being exposed to lab air with approximately 50% relative humidity. This sample, thus, has a low, but finite, degree of hydration. A sample saturated with water was prepared by adding a drop of water to a rotor loosely packed with dry complex salt. The saturated sample was equilibrated at 5 °C for three weeks. Through deconvolution of the resolved MAS 1H spectra, the water contents were determined to 5.7 and 36 water molecules per base pair for the slightly hydrated and the saturated sample, respectively. The amount of water in the saturated sample is in excess of the swelling limit 27 water molecules per base pair.1 Wide-line and Self-diffusion 1H NMR. Wide-line and selfdiffusion 1H experiments were performed on a Bruker DMX200 spectrometer with a Bruker DIFF-25 gradient probe. The 1H resonance frequency for this system is 200 MHz. Used throughout were 4-µs 90° pulses and 4.5-µs receiver dead time. For the CTA hexagonal phase and the DNA-CTA sample with excess D2O, the spectral width was 30 kHz. Free induction decays were recorded as a function of relative humidity and temperature with a dwell time of 1 µs using a fast analog-todigital converter. There were 64 scans accumulated with a recycle delay of 2 s. The 1H background signal from the probe is negligible. The temperature was controlled with an accuracy of 0.5 °C. Self-diffusion measurements were performed with the pulsed-field-gradient stimulated-echo pulse sequence using 1.5-ms gradient pulses and 22.1-ms effective diffusion time. The gradients were incremented in a linear sequence up to 9.6 T/m. The self-diffusion coefficients were evaluated using standard methods.16,17 Magic Angle Spinning. MAS experiments were carried out at 15 °C on a Chemagnetics (Varian) CMX Infinity 500 spectrometer equipped with a 3.2 mm triple-resonance Varian T3 probe. The Larmor frequencies were approximately 500 and 125 MHz for 1H and 13C, respectively. The 1H spectra were recorded as a function of spinning rate using a spectral width of 100 kHz. The 13C spectra with the spectral width 200 kHz were recorded using the wide-line separation (WISE) pulse sequence18 at a 4.2 kHz spinning speed. During 13C detection, 42 kHz 1H dipolar decoupling was applied. For cross polarization, 3-ms contact times were used. Because of the great differences in line width between the different components of the system, the WISE experiment was not performed in the standard way with a linear sequence of 1H evolution times and
J. Phys. Chem. B, Vol. 108, No. 39, 2004 15393 a 2-D Fourier transform analysis. Instead, the 1H evolution times were judiciously chosen to observe both the rapid decay of signal from the solid DNA on the µs time-scale and the slower decay of the liquidlike CTA signal on the ms time-scale. The 13C dimension was evaluated with a standard Fourier transform while the 1H dimension was evaluated by fitting an exponential, for the DNA, or an exponentially damped cosine function, for the CTA, to each peak in the series of 13C spectra. In this way, 1H decay rates and chemical shifts could be estimated using only a fraction of the experimental time needed for a proper 2-D Fourier transform analysis. Still, each WISE experiment required close to 24 h of experimental time with 1000 transients accumulated for each of the 40 1H evolution times. Results and Discussion DNA-CTA Wet and Dry. The DNA-CTA complex is insoluble in water. However, it can absorb reasonable amounts of water under high relative humidity. We have recently measured the sorption isotherm for the DNA-CTA complex.1 As the relative humidity increases from zero, there is a gradual uptake of water. At saturated conditions, the water uptake amounts to 27 ( 1 water molecules per base pair, which corresponds to around seven water molecules per charge. The calorimetric investigations revealed that the sorption enthalpy was negative at low water contents, but it turned distinctly positive at around seven water molecules per base pair and remained so up to the swelling limit. This observation implies that the water uptake is driven by entropy effects except at the lowest water contents. Based on these thermodynamic observations, it was tentatively concluded that the addition of water allows for structural rearrangements in the packing of the hydrocarbon chains of the amphiphile. However, it is notoriously difficult to make definitive molecular interpretations of thermodynamic data for molecularly complex systems and the NMR studies presented in this paper were made in order to monitor the properties of the system on a more direct molecular level. Static 1H Spectra. The width of a 1H resonance line is a sensitive measure of the molecular mobility. For two protons of a CH2 group the maximum dipolar coupling amounts to 60 kHz; for a system that is completely static on the NMR time scale one would observe 1H NMR signals with a width of this magnitude. As the mobility of a sample increases, the dipolar interactions are averaged by motion, and the resonance lines become less broad. For nonviscous liquids, the lines are typically on the order of 1 Hz, the minimum width being limited by the homogeneity of the main magnetic field. Motion on a wide range of length and time scales contributes to the averaging of the dipolar interactions for molecules in supramolecular assemblies. These motions are, for example, conformational rearrangements, rotation of a molecule around its principal axis, lateral molecular diffusion along the aggregate, and rotation of the entire aggregate. Other factors that lead to line broadening are chemical shift anisotropy and susceptibility anisotropy. The former is less important for 1H while the latter may be significant whenever the sample consists of domains with different properties, such as water and oil domains in a liquid crystal or air inclusions in a powder sample. Chemical exchange can be an important factor, especially for water, which may exchange protons with hydroxyl- and amino groups located on rigid macromolecules. In conclusion, wide-line 1H NMR is a sensitive method to detect whether a sample is “mobile” or “rigid” in a general sense, while an accurate estimate of the mobility in terms of correlation times for various modes of motion requires fairly advanced modeling. In Figure 1, we show 1H NMR spectra for the DNA-CTA complex in the dry state (a) and in excess of D2O (b). The
15394 J. Phys. Chem. B, Vol. 108, No. 39, 2004
Figure 1. 1H NMR spectra at 200 MHz for (a) DNA-CTA complex dry, (b) in excess of D2O, and (c) a hexagonal phase of infinite cylinders of CTAB. The scale of the frequency axis is the same for all spectra.
spectra are rather different, which confirms the previous conclusion that structural and/or dynamic changes occur in the swelling process. As a reference, the figure also includes the 1H spectrum for CTAB cylinders arranged in a hexagonal liquid crystalline structure (c), which is analogous to the spectra obtained by Ulmius et al.19 All three spectra in Figure 1 are broad because of finite dipole-dipole couplings. One peak from residual 1H in the D2O and one peak each for the CTA headgroup and hydrocarbon tail can barely be discerned for the hydrated samples. For the completely dry sample, no individual peaks can be resolved. Although broad enough to obscure the chemical shift, the widths of the peaks are still just a fraction of the width that would be obtained for a completely solid sample. For packed alkyl chains, there is normally some internal motion at room temperature that is fast enough to provide some degree of motional narrowing. In fact, the spectrum of the dry complex in Figure 1a resembles the one observed for phospholipids in the lamellar gel state, where the main averaging is due to a rotation around the molecular axis of practically straight alkyl chains.20 We conclude that, even in the dry state of the complex, there are some molecular motions of the chains on a time scale faster than τ < 1/(2πδυ) where δυ is the line width at half-height. In interpreting the spectrum for the fully hydrated sample, we first note that the signal is slightly narrower than that of the liquid crystalline sample of the same amphiphile. There is also a more subtle difference in that the band shapes are different. For the liquid crystal, the signal has the characteristic “superLorentzian” band shape. The source of this shape is a superposition of signals of identical shape but with the width varying as (3cos2θ - 1) where θ is the angle between the alkyl chains’ director and the magnetic field.21 This occurs for a locally ordered system with sufficiently rapid diffusion to average all intermolecular dipolar couplings to zero. We do not observe this shape for the hydrated complex, indicating that there are qualitative differences between the molecular motions in the cylindrical aggregates of the liquid crystal and in the aggregates of the hydrated complex. The smaller overall width of the spectrum for the complex strongly indicates that the amplitude of the molecular motions is such that there is stronger reduction of the intermolecular dipolar couplings than in the liquid crystal. The absence of the “superLorentzian” shape additionally indicates that either translational motion is slow with respect to averaging intermolecular couplings or, alternatively, that the intramolecular couplings are averaged in an environment with less than threefold symmetry. That diffusion is slow is further confirmed by direct measurements giving an upper limit to the self-diffusion coefficient of the surfactant cation of D < 10-13
Leal et al. m2/s. This is in contrast to the values around 10-11 m2/s found for single chain surfactants in liquid crystalline systems.22 Magic Angle Spinning 1H Spectra. Dipolar couplings and susceptibility anisotropy both scale as (3cos2θ-1). By rotating the sample around an axis inclined at 54.7° (the magic angle) to the direction of the magnetic field, the broadening caused by these interactions can be decreased or removed. Significant narrowing occurs when the frequency of rotation exceeds the strength of the interaction (in frequency units). With current MAS equipment, spinning speeds in excess of 20 kHz are routinely available. While this is not sufficient to average the 1H-1H dipolar couplings in solid samples, broadening caused by susceptibility anisotropy, chemical shift anisotropy, small heteronuclear dipolar couplings, and partially motionally averaged 1H-1H dipolar interactions can be removed. In the framework of amphiphile phase behavior studies, MAS experiments have been successfully used to study cubic liquid crystalline phases23 as well as structure and conformational exchange of molecules in lamellar phases.24-26 Solid-state NMR has been also used to study the molecular conformation of different forms of DNA.27 1H spectra for a slightly hydrated and a saturated DNACTA sample as a function of spinning speed are shown in Figure 2. The peaks originate almost exclusively from water and CTA. The signals from DNA are about 10 times broader than the spectral window shown in Figure 1, as is verified in the 13C experiments discussed below. Both static spectra are broad and featureless. For the wet sample, the peaks are narrowed even with slow spinning at 0.5 kHz, indicating that this broadening is primarily caused by the susceptibility difference between the air and the sample grains. The spectrum obtained at fast spinning is quite similar to the spectrum for a micellar solution of CTA. For the dry sample, the peaks are gradually narrowed as the spinning rate is increased. The water peak is already affected at 0.5 kHz, the main methylene peak from the CTA hydrocarbon tail at 3 kHz, and the methyl peak from the CTA headgroup at 10 kHz. The difference between the tail and the headgroup shows that the interiors of the CTA aggregates are more mobile than the exteriors. This interpretation is confirmed by spinecho experiments performed at the highest spinning rate. For the wet sample, the sequence of relative mobility is reversed, with the exterior being the most mobile. These observations imply that the motions of the headgroup are quenched by the proximity of oppositely charged DNA at low hydration. 1H-13C WISE. High-resolution 13C spectra for solid samples can be recorded using CP-MAS. Disordered solids yield broadened peaks because of the sensitivity of the 13C chemical shift to the molecular conformation. By inserting a variable delay between the initial 1H excitation pulse and the onset of CP, the 1H evolution can be detected as an amplitude modulation of the 13C peaks. In this way, high resolution in the 13C dimension is combined with information about mobility in the 1H dimension. These facts form the basis for the WISE experiment.18 13C spectra for a slightly hydrated sample are displayed in Figure 3. Apart from a multitude of narrow peaks originating from the CTA,28 somewhat broadened peaks from the DNA sugars and very broad peaks from the DNA bases are visible.29 The very broad 13C peaks from the DNA bases suggest that these have a low degree of order. In Figure 4, we show the 13C peak area as a function of 1H evolution time in the WISE experiment for the major methylene peak from the CTA hydrocarbon tail, the methylene groups of the CTA headgroup, and the major DNA sugar peak in the slightly hydrated sample. The DNA peaks disappear after a few
Molecular Mobility in a DNA-Amphiphile Complex
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Figure 2. 1H NMR at 500 MHz spectra for slightly hydrated (a) and saturated (b) DNA-CTA as a function of magic angle spinning rate (0, 0.5, 3, and 10 kHz from bottom to top). The major peaks are water at 5 ppm, methyl groups of the CTA headgroup at 3.5 ppm, and methylene groups of the CTA hydrocarbon tail at 1.4 ppm. The spectra are normalized with respect to the water peak. Several spinning sidebands are visible at a 0.5 kHz spinning rate.
Figure 3. 13C NMR spectra for slightly hydrated DNA-CTA obtained with cross polarization from 1H at a magic angle spinning rate of 4.2 kHz. The narrow peaks originate from CTA. Broader peaks from the sugar groups of the DNA can also be seen, three between 60 and 80 ppm and one at 38 ppm. Very broad peaks from the disordered DNA bases are barely visible between 100 and 170 ppm.
Figure 4. 13C peak area as a function of 1H evolution time in the WISE experiment for the major methylene peak from the CTA hydrocarbon tail (circles), the methyl groups of the CTA headgroup (squares), and the major DNA sugar peak (triangles). The lines represent a fit of an exponential (DNA) or an exponentially damped cosine function (CTA) to the data. The fit yields the decay time constants of 19 µs for the DNA sugar, and 0.25 and 0.48 ms for the CTA headgroup and hydrocarbon tail, respectively.
tens of microseconds of 1H evolution, which implies that the DNA is rigid. Fitting an exponential to the decay of the signal yields a relaxation time of 19 µs. The evolution of the CTA peaks was analyzed with an exponentially damped cosine function, yielding relaxation times between 0.25 and 0.85 ms. This observation clearly indicates that even for water contents as low as 1-2 water molecules per charge, CTA is mobile. A similar analysis for the saturated sample yielded relaxation times of 4.7 ms for the CTA headgroup and 1 ms for the CTA
hydrocarbon tail. The order of relative mobility for the different parts of the CTA molecule is the same as that obtained from the experiments with variable spinning rate and spin-echo during MAS. No significant changes of the DNA could be detected upon hydration. Free Induction Decays at Different Water Contents and Temperatures. Wide-line 1H experiments were performed to study the mobility as a function of temperature and hydration level. Although the amount of information is limited in comparison to the WISE experiment, wide-line 1H is useful for the study of gradual changes, because of the relative speed at which the experiment can be performed (2 min for wide-line 1H vs 24 h for WISE). Since no peaks can be resolved in the frequency-domain 1H spectrum, we find it more informative to show the time domain signal, the free induction decay (FID), which upon Fourier transformation gives the spectrum. Figure 5 shows the FID for the DNA-CTA complex equilibrated at (a) different water contents and (b) different temperatures. The slowest decaying component is due to water protons and the relative weight of this signal decreases as the water content decreases. That the slowly decaying component originates from water was checked by repeating the experiment using D2O. In Figure 6 it is shown that, when D2O is used, the slowly decaying component is not present and the first component decays with essentially the same rate as when normal water is used. The DNA signal decays within a few tens of microseconds as was shown with the WISE experiment. From this, we conclude that the change of the dynamic state of the surfactant can be followed using the component of the FID with a decay time of about 0.5 ms. From Figure 5a it is evident that the signal from the alkyl chain protons decays faster the lower the water content, but there is a particularity drastic change of the signal when the water content changes from 5 to 7 water molecules (relative humidities from 53% to 68%). Such a drastic change in the spectral characteristics can reflect a structural transition in the sample. However, the calorimetric data of Leal et al.1 do not reveal any anomalies in the thermodynamic parameters in this concentration range. An alternative interpretation of the source of the spectral changes is obtained by noting that the width of the spectrum is determined by both the amplitude of alkyl chain motions and the rate of these motions. There is no reason to assume that the average conformation of the chains becomes more ordered as the water content is decreased. It is more likely that the observed differences are caused by changes in the rate of conformational transitions. Whether or not motional narrowing occurs is determined by the product of the dipolar interaction
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Leal et al.
Figure 7. Water self-diffusion coefficient as a function of water content within the DNA-CTA complex.
Figure 5. 1H Free Induction Decays obtained at 200 MHz for a DNACTA complex (a) equilibrated at different water contents; from bottom to top water molecules per base pair/relatiVe humidity (%): 0/0, 2.3/ 25, 2.8/30, 3/32, 3.6/43, 5/53, 7/68, 8.7/75, 14.4/88 and (b) equilibrated at different temperatures; from bottom to top °C: -20, -10, 0, 5, 10, 15, 20, 25. The water content of this sample was 8.7 water molecules per base pair (75% relatiVe humidity).
Figure 6. 1H Free Induction Decays obtained at 200 MHz for a DNACTA equilibrated at 43% relative humidity using D2O (grey line) and H2O (black line). The second component is missing when D2O is used and the first component shows essentially the same decay.
(in frequency units) and the characteristic time for the conformational change. It is then conceivable that the relatively large spectral changes that are observed going on from 5 to 7 water
molecules (relative humidities from 53% to 68%) are caused by the fact that the characteristic times for some crucial conformational transitions change in such a way that one goes from static couplings at the lower water content to motional narrowing at the higher one. Based purely on the NMR observations, it is difficult to distinguish between the two alternative interpretations. But, if one also considers the calorimetric measurements on the same system, it appears as the more likely alternative that there is a gradual slowing down of the transitions between the different disordered conformations of the alkyl chain of the surfactant as the water content is decreased. To further investigate the possibility of a surfactant liquidcrystal-to-gel transition, a sample equilibrated to 75% relative humidity (8.7 water molecules per base pair) was studied for a range (-20 to +25 °C) of temperatures. The resulting FIDs are shown in Figure 5b. The decay of the signal from the surfactant is gradually decreasing and there is no sharp transition. The observation further supports the view that in the complex the self-assembled surfactant aggregate is strongly distorted by the very strong electrostatic interactions with the DNA. This enforces an irregular packing of the alkyl chains and there is no basis for a sharp cooperative liquid-solid transition. It appears that there is a large disorder in the chain packing at all water contents, but the time scales for rearrangements decreases with decreasing water content and decreasing temperature. Water Self-diffusion. The dynamic molecular characteristics of the system were further investigated by measuring the water self-diffusion at varying hydration levels. Although the system is locally anisotropic, the decay of the echo amplitude was consistent with a uniform Gaussian diffusion. Since the water is diffusing relatively rapidly, the experiment registers molecular translational displacements over several microns, and the finding of a Gaussian diffusion indicates that the sample is isotropic on the micron length-scale. Figure 7 shows the measured water self-diffusion coefficient in the DNA-CTA complex as a function of water content. The diffusion coefficient increases with increasing water content. For nearly full hydration of the complex (25 water molecules per base pair) the observed diffusion coefficient is approximately 2 × 10-10 m2/s. The diffusion occurs preferentially along the axes of the DNA and CTA cylinders and the measured value can, to a reasonable approximation, be multiplied by a factor of 3 to give D (lateral) ≈ 6 × 10-10 m2/s, considering the local characteristics of the translational mobility. In pure water DH2O (bulk) ) 2.26 × 10-9 m2/s at 25 °C, which is only a factor of 4 times higher. Clearly, even in the very confined environment in the complex, water molecules are very mobile and they are not fully trapped by the discrete charges of the system. As the water content decreases, the diffusion coefficient also decreases. However,
Molecular Mobility in a DNA-Amphiphile Complex
J. Phys. Chem. B, Vol. 108, No. 39, 2004 15397 deswelling, there is a high degree of chain disorder but the dynamics of conformational transitions are gradually slower the lower the water content. The diffusional motion of the CTA molecules is immeasurably slow even at the highest water contents, indicating that the CTA molecules are pinned to the anionic sites of the DNA double helix. This behavior is in sharp contrast to that of the water, for which we measure a reasonably fast diffusion even down to as low a concentration as five water molecules per base pair.
Figure 8. Suggested DNA-CTA packing for an electroneutral complex consisting of a hexagonal arrangement of DNA helices (darker circles) around the central CTA distorted cylinder. From Leal et al.1
even at five water molecules per base pair, corresponding to just over one water molecule per charge, the diffusion is still as rapid as 1 × 10-11m2/s (and locally almost three times higher). In contrast to the water, the CTA self-diffusion is smaller than 10-13 m2/s. This indicates that the CTA cations are interacting so strongly with the DNA phosphate anion groups that they become dynamically pinned. Conclusions NMR provides information on the molecular properties of the CTA cation and the water in the CTA-DNA complex. Combined with the results from a previous calorimetric characterization of the thermodynamic properties of the same system, a coherent picture emerges. The complex is formed due to the strong electrostatic attraction between the negatively charged DNA double helix and the positive CTA cylindrical aggregates. To obtain charge matching at short range, local deformations are necessary and at some point, one obtains a balance between the gain in electrostatic interactions and the free energy cost of aggregate deformation. This occurs at 27 water molecules per base pair for an aggregate in equilibrium with pure water. From the point of view of the surfactant aggregate, the deformations can be seen as of two different types. To obtain charge matching, the linear charge density of the basically cylindrical aggregate has to match that of the corresponding DNA double helices. For a structure as illustrated in Figure 8 where there is a 3:1 DNA-helix-to-CTA-cylinder ratio, one expects an expansion of the CTA cylinder from the optimal area per surfactant. This should then also result in an increase of conformational disorder of the alkyl chains. In addition to the longitudinal expansion, there is always, in principle, a distortion of sixfold symmetry of the basically cylindrical aggregate packed in a hexagonal phase. As the water content decreases, this distortion has to increase in amplitude and it is our understanding that this distortion provides the swelling pressure, or repulsive force, ensuring that the water uptake is up to 27 water molecules. The present investigation shows that under these constraints, due to the deformation, there is no cooperative transition from liquidlike to solidlike behavior of the alkyl chains. Throughout the
Acknowledgment. This work was supported by the Swedish Research Council for Engineering Sciences (C.L. and H.W.) and the Office of Science, Basic Energy Sciences, U.S. Department of Energy under Contract No. DE-AC03-76SF00098 (D.T. and R.W.M.). R.W.M. acknowledges the Department of the Army, contract DAMD 17-03-1-0476, for support. D.T. is financed by a postdoctoral fellowship grant from the Swedish Research Council. The authors acknowledge Prof. Alex Pines for helpful discussions. QOTSA is acknowledged for insightful inspirations (C.L. and D.T.). References and Notes (1) Leal, C.; Wadso¨, L.; Olofsson, G.; Miguel, M.; Wennerstro¨m, H. J. Phys. Chem. B 2004, 108, 3044. (2) Koltover, I.; Salditt, T.; Ra¨dler, J. O.; Safinya, C. R. Science 1998, 281, 78. (3) Ra¨dler, J. O.; Koltover, I.; Salditt, T.; Safinya, C. R. Science 1997, 275, 810. (4) Mel’nikov, S.; Sergeyev, V. G.; Yoshikawa, K.; Takahashi, H.; Hatta, I. J. Chem. Phys. 1997, 107, 6917. (5) Mel’nikov, S. M.; Sergeev, V. G.; Yoshikawa, K. J. Am. Chem. Soc. 1995, 117, 9951. (6) Felgner, P. L.; Gadek, T. R.; Holm, M.; Roman, R.; Chan, H. W.; Wenz, M.; Northrop, J. P.; Ringold, G. M.; Danielsson, M. Proc. Natl. Acad. Sci. U.S.A. 1987, 84, 7413. (7) Felgner, P. L. AdV. Drug DeliVery Res. 1990, 5, 163. (8) Zantl, R.; Artzner, F.; Rapp, G.; Ra¨dler, J. O. Europhys. Lett. 1998, 45, 90. (9) Zantl, R.; Baicu, L.; Artzner, F.; Sprenger, I.; Rapp, G.; Ra¨dler, J. O. J. Phys. Chem. B 1999, 103, 10300. (10) McManus, J. J.; Radler, J. O.; Dawson, K. A. Langmuir 2003, 19, 9630. (11) Pohle, W.; Selle, C.; Gauger, D. R.; Zantl, R.; Artzner, F.; Ra¨dler, J. O. Phys. Chem. Chem. Phys. 2000, 2, 4642. (12) Lindman, B.; So¨derman, O.; Wennerstro¨m, H. NMR Studies of Surfactant Systems. In Surfactant Solutions. New Methods of InVestigation; Zana, R., Ed.; Marcel Dekker: New York, 1987; p 295. (13) Pines, A.; Waugh, J. S.; Gibby, M. G. J. Chem. Phys. 1972, 56, 1776. (14) Andrew, E. R.; Bradbury, A.; Eades, R. G. Nature 1958, 182, 1659. (15) Saenger, W. Principles of Nucleic Acid Structure; SpringerVerlag: New York, 1984. (16) So¨derman, O.; Stilbs, P. Prog. Nucl. Magn. Reson. Spectrosc. 1994, 26, 445. (17) Johnson, C. S., Jr. Prog. Nucl. Magn. Reson. Spectrosc. 1999, 34, 203. (18) Schmidt-Rohr, K.; Clauss, J.; Spiess, H. W. Macromolecules 1992, 25, 3273. (19) Ulmius, J.; Wennerstro¨m, H. J. Magn. Reson. 1977, 28, 309. (20) Ulmius, J.; Wennerstro¨m, H.; Lindblom, G.; Arvidson, G. Biochim. Biophys. Acta 1975, 389, 197. (21) Wennerstro¨m, H. Chem. Phys. Lett. 1973, 18, 41. (22) Lindblom, G.; Wennerstro¨m, H. Biophys. Chem. 1977, 6, 167. (23) Pampel, A.; Strandberg, E.; Lindblom, G.; Volke, F. Chem. Phys. Lett. 1998, 287, 468. (24) Lindblom, G. Curr. Opin. Colloid Interface Sci. 1996, 1, 287. (25) Bystro¨m, T.; Grobner, G.; Lindblom, G. Colloids Surf., A 2003, 228, 37. (26) Strandberg, E.; Sparrman, T.; Lindblom, G. AdV. Colloid Interface Sci. 2001, 89, 239. (27) van Dam, L.; Levitt, M. H. J. Mol. Biol. 2000, 304, 541. (28) Alonso, B.; Massiot, D. J. Magn. Reson. 2003, 163, 347. (29) Santos, R. A.; Tang, P.; Harbison, G. S. Biochemistry 1989, 28, 9372.
