Vibrational Dynamics and Couplings of the Hydrated RNA Backbone

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Cite This: J. Phys. Chem. Lett. 2018, 9, 583−587

Vibrational Dynamics and Couplings of the Hydrated RNA Backbone: A Two-Dimensional Infrared Study Eva M. Bruening, Jakob Schauss, Torsten Siebert, Benjamin P. Fingerhut, and Thomas Elsaesser* Max-Born-Institut für Nichtlineare Optik und Kurzzeitspektroskopie, Max-Born-Str. 2a, D-12489 Berlin, Germany S Supporting Information *

ABSTRACT: The equilibrium structure of the RNA sugar−phosphate backbone and its hydration shell is distinctly different from hydrated DNA. Applying femtosecond twodimensional infrared (2D-IR) spectroscopy in a range from 950 to 1300 cm−1, we elucidate the character, dynamics, and couplings of backbone modes of a double-stranded RNA Ahelix geometry in its aqueous environment. The 2D-IR spectra display a greater number of backbone modes than for DNA, with distinctly different lineshapes of diagonal peaks. Phosphate−ribose interactions and local hydration structures are reflected in the complex coupling pattern of RNA modes. Interactions with the fluctuating water shell give rise to spectral diffusion on a 300 fs time scale, leading to a quasi-homogeneous line shape of the symmetric (PO2)− stretching mode of the strongly hydrated phosphate groups. The RNA results are benchmarked by 2D-IR spectra of DNA oligomers in water and analyzed by molecular dynamics and quantum mechanical molecular mechanics simulations.

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tional modes are to some extent contradictory and the impact of hydration geometries on the mode pattern has remained unknown.9−12 At ambient temperature, both the RNA A-helix and the surrounding water shell undergo structural fluctuations on a multitude of time scales, the fastest in the femtosecond range. The character of these fluctuations, the strength of the underlying interactions, and their consequences for the functional properties of RNA are not well-understood. Vibrational excitations of the helix backbone are attractive, noninvasive, and local probes of such processes, as has recently been demonstrated by two-dimensional infrared (2D-IR) spectroscopy of hydrated DNA.13−15 Here, we apply 2D-IR spectroscopy to analyze the character and couplings of RNA backbone modes in the 950−1300 cm−1 range, to map RNA− water interactions, and to determine the time scale of fluctuations as well as the dynamic heterogeneity of hydration sites in the backbone. Benchmarking the results by comparative measurements with hydrated DNA reveals striking differences between the two systems. Double-stranded RNA oligomers containing 23 alternating adenine-uracil (A-U) base pairs in Watson−Crick geometry are studied in an aqueous environment with more than 190 water molecules per base pair (c(RNA) = 10−2 M). Under such conditions of full hydration, RNA forms an A-helix structure. The DNA B-helix reference structure consists of 23 alternating adenine-thymine (A-T) base pairs in Watson−Crick geometry and is dissolved in H2O with a similar concentration. Both samples contain Na+ counterions. In Figure 1, infrared

he interaction of RNA and DNA with water molecules of the surrounding hydration shell plays a decisive role in their macromolecular structure and chemical function.1,2 Timeaveraged equilibrium structures and hydration patterns have been derived from X-ray diffraction data recorded from crystallized samples.3,4 In these studies, the positions of water molecules at the RNA and DNA surfaces have been inferred from the positions of the electron-rich oxygen atoms while the water hydrogens have remained mostly elusive. Such experimental information has been complemented by molecular dynamics (MD) simulations5−8 from which time-averaged structures, radial distribution functions along particular RNAwater and water−water hydrogen bonds, and hydration dynamics have been derived. RNA exists in both single- and double-stranded form, with a secondary structure adapted to its particular biochemical function. For full hydration at physiological temperatures, short double-stranded RNA forms a well-defined A-helix structure with bases paired in a Watson−Crick geometry. Fully hydrated DNA displays a B-type double-helix while Aform double-helices of DNA prevail at low hydration. The sugar−phosphate backbone of RNA contains an OH group attached to the 2′ carbon atom of the ribose rings. This hydroxyl group which is absent in DNA strongly affects the hydration pattern at the RNA surface. The 2′−OH groups induce a regular and well-defined hydration pattern in the minor groove of the RNA helix, as has been discussed in detail in ref 3. Adjacent phosphate groups are bridged by individual water molecules, and the overall number of water molecules in the first hydration layer around RNA is somewhat higher than for DNA.3,5 The presence of the 2′−OH group and the A-helix geometry of RNA result in a vibrational spectrum of backbone modes which is more complex than in DNA.9,10 Because of this increased complexity, assignments of RNA backbone vibra© XXXX American Chemical Society

Received: December 15, 2017 Accepted: January 16, 2018 Published: January 16, 2018 583

DOI: 10.1021/acs.jpclett.7b03314 J. Phys. Chem. Lett. 2018, 9, 583−587

Letter

The Journal of Physical Chemistry Letters

Figure 1. Infrared absorption spectra of backbone modes of (a) a double-stranded DNA helix containing 23 adenine-thymine (A-T) base pairs and (b) a double-stranded RNA helix containing 23 adenine-uracil (A-U) base pairs, both in an aqueous environment under conditions of full hydration. The RNA spectrum displays additional bands compared to DNA. The letters in panels a and b refer to the assignments made in the text. Insets: molecular structure of a backbone segment of DNA in B-helix geometry and RNA in A-helix geometry. (c) Infrared spectra from QM/MM normal mode simulations with RNA backbone geometries taken from molecular dynamics simulations. The dashed line gives the average over all realizations; the solid lines represent spectra for hydration patterns without counterions in the first water layer with νP1 < 1250 cm−1 (blue), 1240 < νP1 < 1250 cm−1 (red), and νP1 < 1240 cm−1 (green).

absorption spectra of (a) DNA and (b) RNA are presented. The absorbance A = −log(T0) normalized to the peak value of the asymmetric (PO2)− stretching band is plotted as a function of wavenumber (T0: sample transmission). The broad and structureless background absorption from water librations16 was subtracted in both spectra. The reference spectrum of the DNA B-helix (Figure 1a) comprises seven main components, the asymmetric and symmetric (PO2)− stretching bands P1 and P2 at 1225 and 1088 cm−1, the diester linkage modes L1 and L2 at 1070 and 1054 cm−1, the furanose ring mode R1 at 1016 cm−1, and the ribose main chain and diester linkage modes R2 and L3 giving rise to the peak at 970 cm−1.13,17,18 The spectrum of the RNA A-helix (Figure 1b) displays P1 and P2 bands at 1245 and 1087 cm−1, the latter with a shoulder at 1100 cm−1. The L2 band occurs in the range around 1060 cm−1, similar to A-form DNA.9−11,19,20 The band at 916 cm−1 is due to a ribose mode,21 and the lines at 970 and 995 cm−1 have tentatively been assigned to ribose−phosphate main chain vibrations.9 The assignments of the characteristic RNA bands at 1120 and 1220 cm−1 and shoulders around 1100 and 1135 cm−1 have remained speculative and will be discussed below. Figure 2a,b shows experimental 2D-IR spectra of RNA in different, partially overlapping spectral ranges. The absorptive 2D signal recorded at a waiting time of T = 250 fs and

Figure 2. Two-dimensional infrared (2D-IR) spectra of RNA backbone vibrations measured in a frequency range (a) from 940 to 1150 cm−1 and (b) from 1030 to 1280 cm−1. The absorptive 2D signal is plotted as a function of excitation frequency ν1 and detection frequency ν3. The signal amplitudes are normalized to the maximum positive signal in each spectrum. Yellow-red contours represent positive signals, while blue contours are negative signals. The steps in signal amplitude are 5% between adjacent contour lines. The positive components of the diagonal peaks are due to bleaching and stimulated emission on the fundamental v = 0 to 1 transition of the respective vibration, the negative components are caused by the anharmonically red-shifted transient v = 1 to 2 absorption. (c) Simulated 2D-IR spectra. The parameters of the calculation are summarized in Table 1.

normalized to the maximum positive signal is plotted as a function of excitation (ν1) and detection frequency (ν3). The 2D-IR spectrum in Figure 2a displays a series of diagonal peaks, the strongest one at the frequency position of the symmetric stretching mode (P2) of the (PO2)− units at 1087 cm−1. The 2D signal strength S2D ∝ μ4 is proportional to the fourth power of the vibrational transition dipole μ while A ∝ μ2 holds for the linear absorbance A. As a result, the diagonal peaks in the 2D spectrum show a stronger contrast and a better spectral separation than the bands in the linear absorption spectrum (Figure 1b). At (1102, 1102) cm−1, there is a distinct diagonal 584


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at 1247 and 1220 cm−1 is minor while the latter mode strongly couples to the 1120 cm−1 mode (cf. Figure 2b, (1220, 1120) and (1120, 1220) cm−1 crosspeaks) and the P2 mode. The 2D lineshapes of the P2 peaks of RNA and DNA and the adjacent modes of RNA are presented on a magnified frequency scale in Figure 3. The lineshapes of the P2 mode of RNA (Figure 3a) exhibit an orientation parallel to the ν1 axis, revealing a predominant homogeneous broadening. In contrast, the P2 line shape of DNA (Figure 3b) is tilted with respect to the ν1 axis, suggesting an inhomogeneous contribution to the spectral envelope. The RNA diagonal and cross peaks around ν3 = 1103 and 1130 cm−1 are absent in the DNA spectrum. The envelope of the RNA diagonal peak at (1130, 1130) cm−1 is asymmetrically extended toward higher frequencies, pointing to a substructure. Cuts along the frequency diagonal ν1 = ν3 (Figure S3 in the Supporting Information) confirm the asymmetric shape of this peak. The presence of different subcomponents is supported by the positions of the cross peaks at (1135, 1087) and (1090, 1135) cm−1, which are shifted with respect to the maximum position of the diagonal peak. As a function of waiting time T, the amplitudes of all 2D peaks decay on a 1−2 picosecond time scale (cf. Figure S2) without a noticeable reshaping of their spectral envelopes. Independent pump−probe measurements were performed to determine the v = 1 decay times of the different modes. Experimental results are summarized in the Supporting Information, and the extracted decay times are listed in Table 1. Quantum mechanical molecular mechanics (QM/MM) normal mode analyses were performed for an alternating 23mer A-U RNA helix along snapshots of MD trajectories (3·0.2 μs = 0.6 μs total simulation time) to assign the different backbone normal modes to structural units of the backbone (details of the theoretical approach are presented in the Supporting Information). Calculated linear infrared spectra are presented in Figure 1c and Figure S7, and the predominant character of the respective mode displacements is summarized in Table 1 and Table S1. Solid lines in Figure 1c represent spectra with a neat water solvation environment that is obtained by frequency filtering of P1 modes (νP1 < 1250 cm−1, cf. Supporting Information for details), while the dashed line represents the average over all realizations along the RNA helix including those with counterions in close vicinity, like, e.g., within the first hydration layer (Figure S6). According to our analysis, the prominent doublet above 1200 cm−1 arises from asymmetric (PO2)− stretching vibrations νP1 where the frequency position depends sensibly on the water hydration geometries and the presence of ions (Figures S6 and

Figure 3. Comparison of (a) RNA and (b) DNA 2D-IR spectra in the range around the symmetric (PO2)− stretching signal at ν3 ≈ 1090 cm−1. The RNA spectrum displays a subset of additional diagonal and cross peaks as well as different lineshapes of diagonal peaks.

peak while the linear absorption spectrum just shows a spectral shoulder. Because of the substantially smaller transition dipoles, the diagonal peaks below (ν1, ν3) = (1050, 1050) cm−1 appear as weak features only. The particularly rich cross peak pattern with positive and negative components demonstrates pronounced anharmonic couplings between the backbone modes in a range of ν3 from 1050 to 1150 cm−1. The 2D-IR spectrum in Figure 2b reveals the prominent peak due to the asymmetric (PO2)− stretching mode P1 around (1247, 1247) cm−1 together with a weaker peak around (1220, 1220) cm−1. The diagonal P1 peak at (1247, 1247) cm−1 exhibits an elliptic line shape, pointing to an inhomogeneous broadening contribution. In contrast, the P2 diagonal peak around (1087, 1087) cm−1 is oriented parallel to the ν1 axis, a hallmark of minor inhomogeneous broadening and fast spectral diffusion. The crosspeak pattern in Figure 2b reveals a pronounced coupling between P1 and P2 by the pair of positive and negative cross peak components at (1088, 1253) and (1088, 1226) cm−1. Notably, coupling between the modes Table 1. Backbone Normal Modes of RNAa frequency (cm−1)

anharm. (cm−1)

lifetime (ps)

FFCF Δ1 (cm−1)

FFCF Δ2 (cm−1)

νC−O−C νC2′−OH

asymmetric (PO2) stretch asymmetric (PO2)− stretch ribose C1′−O4′−C4′ stretch C2′−OH stretch

1245 1220 1133 1120

10 8 10 9

0.36 ± 0.1 − 0.8 ± 0.3 1.1 ± 0.2

14.0 (13.0) 8 5.3 8.0

7.0 (9.5) 3.0 4.8 5.3

νL1 νP2 νL2 νR′1 νR′2

linker C−O stretch symmetric (PO2)− stretch linker C−O stretch ribose main chain (C1′−O4′−C4′−C3′ stretch) ribose main chain (C4′−C5′ ethyl stretch)

1102 1087 1062 994 970

11 10 11 10 10

− 1.0 ± 0.2 − − −

7.0 9.0 (13) 13.0 7.4 6.9

5.3 1.1 (7) 9.0 6.4 5.3

mode νP1(1) νP1(2)

character −

The FFCF amplitude values Δ1 and Δ2 in parentheses are derived from a numerical analysis of the 2D-IR spectra of fully hydrated DNA oligomers (23 A-T pairs); for details of the assignment of mode character, see the Supporting Information. a

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mode P2, the fast component of the FFCF strongly dominates, leading to the practically homogeneous 2D line shape. X-ray diffraction studies3 and molecular dynamics simulations5 have suggested that the first hydration layer around RNA contains a somewhat larger number of water molecules in a more ordered arrangement, compared to DNA. In the A-helix geometry of RNA, the distance between neighboring phosphate groups is small enough to be bridged by a single water molecule which forms strong hydrogen bonds with the respective (PO2)− oxygens.3,5,23 The presence of the 2′−OH group of the RNA ribose units not only introduces additional backbone modes but also influences the water structure in the minor and, to lesser extent, the major groove. This behavior has been rationalized with the concept of hydrogen bonded rings of water molecules.3 Both structural features of the RNA backbone reduce the heterogeneity of the first water layer and thus are consistent with the reduced inhomogeneous width of the 2D lineshapes of RNA compared to DNA. The decay time of the fast FFCF component of 300 fs is shorter than the picosecond hydrogen bond lifetimes at the water−RNA interface. A very similar 300 fs decay has been observed with hydrated DNA13,15,24 and phospholipids.25 The slowing of fluctuations is moderate compared to bulk water.26 The fast decay mainly reflects fluctuations of short-range electric fields which originate from the dipolar water molecules in the first and second hydration layer.27 Water molecules in this environment undergo subpicosecond librational motions within their steric constraints set by the RNA or DNA surface, and/or hydrogen bonds between water molecules in a geometry distinctly different from bulk water.2,3,5 In the DNA B-helix, the methyl group of thymine defines a steric constraint for a near-by water molecule which forms a hydrogen bond with a phosphate oxygen and is considered to be “immobilized” by the methyl group.4,28 The methyl group and resulting steric constraints are absent in the uracil units of RNA. However, the similarity of the fast FFCF decays in RNA and DNA points to a minor role of this structural feature for hydration dynamics on a subpicosecond time scale. The asymmetric (PO2)− stretching mode P1 is particularly sensitive to interactions with the aqueous environment via hydrogen bonds and electronic polarization effects.24,25,27 The P1 bands in both the linear (cf. Figure 1b) and the 2D-IR spectra are split into two subcomponents with maxima at (1220, 1220) and (1247, 1247) cm−1, a behavior markedly different from DNA13 (cf. Figure 1a) and phospholipids.25 The absence of cross peaks between the two components demonstrates their uncoupled nature, thus originating from distinctly different phosphate sites in the backbone. The 1220 cm−1 mode displays prominent cross peaks with the P2 mode at 1087 cm−1 and the ribose C2′−OH and C−O−C modes at 1120 and 1133 cm−1. In contrast, the 1247 cm−1 P1 mode strongly couples to P2, similar to the behavior of B-helix DNA, but shows reduced coupling to the C2′−OH and C−O−C modes of the ribose moiety. The X-ray diffraction data of ref 3 suggest the presence of RNA sites where individual water molecules link the oxygen atom of the ribose 2′−OH group with O3′ and O4′ ribose oxygen atoms and the O2P atom of the phosphate group. Such water molecules may be instrumental in inducing pronounced vibrational couplings between the phosphate mode P1 at 1220 cm−1 and the ribose unit. It should be noted that the limited inhomogeneous broadening of the diagonal peaks indicates a more ordered character of the hydration shell structure around RNA, which is

S8). In particular, the latter induces a strong blue-shift of the asymmetric stretching frequency to values above 1280 cm−1. In contrast, the frequency positions of all other vibrations, including the symmetric (PO2)− stretch vibration, are much less sensitive to the detailed solvation structure. The bands at 1120 and 1135 cm−1 originate from C2′−OH modes involving the OH group in the 2′ position, and additional contributions from C−O−C stretching modes involving the C4′, O4′, and C1′ atoms of the ribose units and are absent in the DNA infrared spectrum (Figure 1a). The linker C−O stretching modes νL1/L2 connecting the phosphate groups with the ribose backbone are located at 1062 and 1102 cm−1, both gaining oscillator strength from coupling to the symmetric (PO2)− stretching mode νP2. The relative absorption strengths of such coupled modes are sensitive to the hydration structure, as is suggested by the spectra in Figure 1c (solid lines). The 2D-IR spectra were analyzed by theoretical simulations combining a density matrix approach for calculating the nonlinear vibrational response functions in third order22 and a Kubo ansatz for the frequency fluctuation correlation function (FFCF) of the different vibrations. Details of this treatment are presented in the Supporting Information and in ref 13 (supplement). The Kubo ansatz for the FFCF is composed of two exponentially decaying terms with amplitudes Δ1 and Δ2 and decay times τ1 = 300 fs and τ2 = 50 ps, similar to our previous studies of DNA hydration dynamics.13−15 The fast component gives rise to spectral diffusion of vibrational frequencies, whereas the slow component with a decay time much longer than the 0.36−1.1 ps vibrational lifetimes of the backbone modes (Table 1) causes quasi-static inhomogeneous broadening. The microscopic origin of fast spectral diffusion is the fluctuating electric force the hydration shell exerts on the backbone oscillators. Inhomogeneous broadening reflects the structural heterogeneity of hydrated RNA, in particular variations in the local water geometries and hydrogen bond strengths and in the presence of counterions. In the simulations, the time constants τ1 and τ2 were fixed and the amplitudes Δ1 and Δ2 were adjusted to account for the lineshapes of the different backbone modes. Lifetime broadening of the bands is included using the v = 1 decay times from the pump−probe experiments. To account for the cross peaks in the 2D-IR spectra, all nonzero intermode couplings were simulated with a single coupling strength of 10 cm−1. The calculated 2D-IR spectrum is plotted in Figure 2c, and cuts through this spectrum are presented in Figure S3. The amplitude values Δ1 and Δ2 for each mode are summarized in Table 1. The calculated and the experimental 2D-IR spectra (Figure 2b) are in good agreement, both displaying a substantial variation of the 2D lineshapes of the different diagonal peaks with respect to spectral width and orientation in the 2D frequency plane. This behavior originates mainly from the different amplitudes of the fast and slow component of the FFCF for the individual modes. The amplitudes Δ2 of the slow component cover a wide range from 1 to 10 cm−1 and are somewhat smaller than for the backbone modes of hydrated DNA (cf. Table 1). This fact results in a limited inhomogeneous broadening and a clear separation of most diagonal peaks. The amplitudes Δ1 of the fast correlation decay are between 5 and 14 cm−1 and determine, together with the respective lifetime broadening, the width of the 2D lineshapes in antidiagonal direction. For the symmetric (PO2)− stretch 586


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(9) Tsuboi, M. Application of Infrared Spectroscopy to Structure Studies of Nucleic Acids. Appl. Spectrosc. Rev. 1970, 3, 45−90. (10) Banyay, M.; Sarkar, M.; Gräslund, A. A Library of IR Bands of Nucleic Acids in Solution. Biophys. Chem. 2003, 104, 477−488. (11) Lindqvist, M.; Sarkar, M.; Winqvist, A.; Rozners, E.; Strömberg, R.; Gräslund, A. Optical Spectroscopy Study of the Effects of a Single Deoxyribose Substitution in a Ribose Backbone: Implications in RNARNA Interaction. Biochemistry 2000, 39, 1693−1701. (12) Kolkenbeck, K.; Zundel, G. The Significance of the 2′ OH Group and the Influence of Cations on the Secondary Structure of the RNA Backbone. Biophys. Struct. Mech. 1975, 1, 203−219. (13) Siebert, T.; Guchhait, B.; Liu, Y.; Costard, R.; Elsaesser, T. Anharmonic Backbone Vibrations in Ultrafast Processes at the DNA− Water Interface. J. Phys. Chem. B 2015, 119, 9670−9677. (14) Guchhait, B.; Liu, Y.; Siebert, T.; Elsaesser, T. Ultrafast Vibrational Dynamics of the DNA Backbone at Different Hydration Levels Mapped by Two-Dimensional Infrared Spectroscopy. Struct. Dyn. 2016, 3, 043202. (15) Siebert, T.; Guchhait, B.; Liu, Y.; Fingerhut, B. P.; Elsaesser, T. Range, Magnitude, and Ultrafast Dynamics of Electric Fields at the Hydrated DNA Surface. J. Phys. Chem. Lett. 2016, 7, 3131−3136. (16) Bertie, J. E.; Lan, Z. Infrared Intensities of Liquids XX: The Intensity of the OH Stretching Band of Liquid Water Revisited, and the Best Current Values of the Optical Constants of H20(I) at 25 °C between 15,000 and 1 cm−1. Appl. Spectrosc. 1996, 50, 1047−1057. (17) Guan, Y.; Thomas, G. J. Vibrational Analysis of Nucleic Acids. IV. Normal Modes of the DNA Phosphodiester Structure Modeled by Diethyl Phosphate. Biopolymers 1996, 39, 813−835. (18) Guan, Y.; Thomas, G. J. Vibrational Analysis of Nucleic Acids. V. Force Field and Conformation-Dependent Modes of the Phosphodiester Backbone Modeled by Diethyl Phosphate. Biophys. J. 1996, 71, 2802−2814. (19) Liquier, J.; Akhebat, A.; Taillandier, E.; et al. Characterization by FTIR Spectroscopy of the Oligoribonucleotide Duplexes r(A-U)6 and r(A-U)8. Spectrochim. Acta 1991, 47A, 177−186. (20) Brown, E. B.; Peticolas, W. L: Conformational Geometry and Vibrational Frequencies of Nucleic Acid Chains. Biopolymers 1975, 14, 1259−1271. (21) Carmona, P.; Molina, M. Raman and Infrared Spectra of DRibose and D-Ribose 5-Phosphate. J. Raman Spectrosc. 1990, 21, 395− 400. (22) Mukamel, S. Multidimensional Femtosecond Correlation Spectroscopies of Electronic and Vibrational Excitations. Annu. Rev. Phys. Chem. 2000, 51, 691−729. (23) Saenger, W.; Hunter, W. N.; Kennard, O. DNA Conformation is Determined by Economics in the Hydration of Phosphate Groups. Nature 1986, 324, 385−388. (24) Floisand, D. J.; Corcelli, S. Computational Study of Phosphate Vibrations as Reporters of DNA Hydration. J. Phys. Chem. Lett. 2015, 6, 4012−4017. (25) Costard, R.; Heisler, I. A.; Elsaesser, T. Structural Dynamics of Hydrated Phospholipid Surfaces Probed by Ultrafast 2D Spectroscopy of Phosphate Vibrations. J. Phys. Chem. Lett. 2014, 5, 506−511. (26) Kraemer, D.; Cowan, M. L.; Paarmann, A.; Huse, N.; Nibbering, E. T. J.; Elsaesser, T.; Miller, R. J. D. Temperature Dependence of the Two-Dimensional Infrared Spectrum of Liquid H2O. Proc. Natl. Acad. Sci. U. S. A. 2008, 105, 437−442. (27) Fingerhut, B. P.; Costard, R.; Elsaesser, T. Predominance of Short Range Coulomb Forces in Phosphate-Water Interactions − A Theoretical Analysis. J. Chem. Phys. 2016, 145, 115101. (28) Drew, H. R.; Dickerson, R. E. Structure of a B-DNA Dodecamer. III. Geometry of Hydration. J. Mol. Biol. 1981, 151, 535−556.

essential for separating the two P1 components and respective coupling patterns. In conclusion, our 2D-IR study of RNA backbone modes in combination with in-depth theoretical simulations reveals the microscopic normal mode character, the intermode coupling pattern, and the subpicosecond time scale of structure fluctuations in the hydration shell. Key features of the 2D-IR spectra such as the limited inhomogeneous broadening of the 2D envelopes and the cross peak pattern correlate with structural insight from X-ray diffraction data. Compared to DNA, hydrated RNA shows a richer vibrational spectrum and coupling scheme as well as less structural disorder in the first hydration layer. Noteworthy is the distinctly different coupling between motions of the ribose and the phosphate group in DNA and RNA, an aspect that may affect the function of the respective backbone structures. Our results illustrate the potential of 2D-IR spectroscopy for unraveling the interplay of structure and dynamics in complex biomolecular systems.

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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jpclett.7b03314. Experimental methods and data analysis, 2D infrared spectra, pump−probe results, and theoretical methods (PDF)

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AUTHOR INFORMATION

ORCID

Eva M. Bruening: 0000-0002-6041-3638 Benjamin P. Fingerhut: 0000-0002-8532-6899 Thomas Elsaesser: 0000-0003-3056-6665 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS B.P.F. gratefully acknowledges support through the DFG within the Emmy Noether Programme (Grant No. FI 2034/1-1). REFERENCES

(1) Saenger, W. Principles of Nucleic Acid Structure; Springer: Berlin, 1984; Ch. 17. (2) Laage, D.; Elsaesser, T.; Hynes, J. T. Water Dynamics in the Hydration Shells of Biomolecules. Chem. Rev. 2017, 117, 10694− 10725. (3) Egli, M.; Portmann, S.; Usman, N. RNA Hydration: A Detailed Look. Biochemistry 1996, 35, 8489−8494. (4) Vlieghe, D.; Turkenburg, J. P.; van Meervelt, L. B-DNA at Atomic Resolution Reveals Extended Hydration Patterns. Acta Crystallogr., Sect. D: Biol. Crystallogr. 1999, 55, 1495−1502. (5) Auffinger, P.; Westhof, E. Water and Ion Binding Around RNA and DNA (C,G) Oligomers. J. Mol. Biol. 2000, 300, 1113−1131. (6) Duboué-Dijon, E.; Fogarty, A. C.; Hynes, J. T.; Laage, D. Dynamical Disorder in the DNA Hydration Shell. J. Am. Chem. Soc. 2016, 138, 7610−7620. (7) Besseova, I.; Banas, P.; Kuhrova, P.; Kosinova, P.; Otyepka, M.; Sponer, J. Simulations of A-RNA Duplexes. The Effect of Sequence, Solute Force Field, Water Model, and Salt Concentration. J. Phys. Chem. B 2012, 116, 9899−9916. (8) Dans, P. D.; Danilane, L.; Ivani, I.; Drasata, T.; Lankas, F.; Hospital, A.; Walther, J.; Pujagut, R. I.; Battistini, F.; Gelpi, J. L.; Lavery, R.; Orozco, M. Long-Timescale Dynamics of the DrewDickerson Dodecamer. Nucleic Acids Res. 2016, 44, 4052−4066. 587


Vibrational Dynamics and Couplings of the Hydrated RNA Backbone

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