Excited state structure/dynamics

EXCITED-STATE STRUCTURAL DYNAMICS OF NUCLEIC ACIDS AND THEIR COMPONENTS... 

Keywords Excited-State Structural Dynamics, Nucleic Acids, Resonance Raman Spectroscopy,... 

Excited-State Structural Dynamics of Nucleic Acids and Their Components 239... 

A critical pre-requisite to using Raman and resonance Raman spectroscopy to examine the excited-state structural dynamics of nucleic acids and their components, is the determination of the normal modes of vibration for the molecule of interest. The most definitive method for determining the normal modes is exhaustive isotopic substitution, subsequent measurement of the IR and Raman spectra, and computational analysis with the FG method of Wilson, Decius, and Cross [77], Such an analysis is rarely performed presently because of the improvements in accuracy of ab initio and semi-empirical calculations. Ab initio computations have been applied to most of the nucleobases, which will be described in more detail below, resulting in relatively consistent descriptions of the normal modes for the nucleobases. 

Within the separable harmonic approximation, the < f i(t) > and < i i(t) > overlaps are dependent on the semi-classical force the molecule experiences along this vibrational normal mode coordinate in the excited electronic state, i.e. the slope of the excited electronic state potential energy surface along this vibrational normal mode coordinate. Thus, the resonance Raman and absorption cross-sections depend directly on the excited-state structural dynamics, but in different ways mathematically. It is this complementarity that allows us to extract the structural dynamics from a quantitative measure of the absorption spectrum and resonance Raman cross-sections. 

The time correlator method [85, 88, 89] has not been used as frequently as the other methods described here, and has not been used for nucleic acids and their components. It will therefore not be discussed further. The experimental methods for determining absolute Raman and resonance Raman cross-sections have been extensively reviewed [90-93], Similarly, the methods for practical use of the sum-over-states, time-dependent, and transform methods for determining excited-state structural dynamics have been extensively reviewed [78-83],... 

The topic of this review is the excited-state structural dynamics of nucleic acids and their components. As stated above, the excited states of nucleic acid components... 

One of the most interesting nucleobases in which to study the excited-state structural dynamics is thymine, as thymine photoproducts account for >95% of the lesions found in DNA upon either UVB or UVC irradiation [59], Excited-state structural... 

With this normal mode description, then, it is instructive to review the resonance Raman intensity-derived excited-state structural dynamics. The first UV resonance Raman study of thymine was not done until 1994 by Lagant, et al. [113], Although Raman and IR spectra of thymine had been recorded much earlier. Most earlier studies of nucleic acid components focussed on the nucleosides and nucleotides. Indeed, much of the earlier research on nucleic acid components was done by the groups of Peticolas and Spiro, working independently. Spiro focussed more on nucleosides and larger nucleic acid structures (see below), while Peticolas examined the nucleobases initially. Peticolas s approach was to combine ab initio computations of the ground-state and excited-state structures and vibrational frequencies, with... 

The UV resonance Raman spectrum of thymine was revisited in 2007, with a slightly different approach, by Yarasi, et al. [119]. Here, the absolute UV resonance Raman cross-sections of thymine were measured and the time-dependent theory was used to experimentally determine the excited-state structural dynamics of thymine. The results indicated that the initial excited-state structural dynamics of thymine occurred along vibrational modes that are coincident with those expected from the observed photochemistry. The similarity in a DFT calculation of the photodimer transition state structure [29] with that predicted from the UV resonance Raman cross-sections demonstrates that combining experimental and computational techniques can be a powerful approach in elucidating the total excited-state dynamics, electronic and vibrational, of complex systems. 

UV resonance Raman spectra of chemical analogues of the pyrimidine nucleobases have been recently reported. Billinghurst et al. [142] have recently reported the UV resonance Raman intensity-derived excited-state stmctural dynamics of 5-fluorouracil, and have shown them to be essentially identical to those of thymine. In that paper, they also report the UV resonance Raman spectra of 5-chlorouracil, 5-bromouracil, and 5-iodouracil, and show that the spectra all are similar. By extension, they argue that the excited-state stmctural dynamics of the 5-halouracils are all similar to each other and to those of thymine, supporting their model that the excited-state structural dynamics of uracil and thymine nucleobases are dictated by the mass of the substituent at the 5 position. 

Very few reports of the excited-state structural dynamics of the purine nucleobases have appeared in the literature. This lack of research effort is probably due to a number of factors. The primary factor is the lack of photochemistry seen in the purines. Although adenine can form photoadducts with thymine, and this accounts for 0.2% of the photolesions found upon UVC irradiation of DNA [67], the purines appear to be relatively robust to UV irradiation. This lack of photoreactivity is probably due to the aromatic nature of the purine nucleobases. A practical issue with the purine nucleobases is their insolubility in water. While adenine enjoys reasonable solubility, it is almost an order of magnitude lower than that of thymine and uracil, the two most soluble nucleobases [143], Guanine is almost completely insoluble in water at room temperature [143],... 

Nevertheless, a few reports of UV resonance Raman spectra of the purine nucleobases and their derivatives have appeared. Peticolas s group has reported the identification of resonance Raman marker bands of guanine, 9-methylguanine and 9-ethylguanine for DNA conformation [118, 144], In the process of doing that work, very rudimentary excitation profiles were measured, which yielded preliminary structures for two of the ultraviolet excited electronic states. Tsuboi has also performed UV resonance Raman on purine nucleobases in an effort to determine the resonance enhanced vibrational structure [94], Thus far, no excited-state structural dynamics for any of the purine nucleobases have been determined. 

In contrast to the isolated nucleobases, a significant amount of work has been done on the excited-state structural dynamics and UV resonance Raman spectroscopy of the nucleotides. The excited-state structural dynamics of nucleosides have not been determined to date, although some UV resonance Raman spectra of the nucleosides have been measured [139, 145, 146], This work on the nucleosides has been primarily for the application of UV resonance Raman spectroscopy in the determination of ground-state structure, rather than excited-state structural dynamics. This emphasis on the nucleotides is no doubt driven by their greater solubility, as well as their immediate relevance to the nucleic acids. In addition, it was believed that the vast majority of the intensity observed in the UV Raman spectra arose through resonance enhancement of the nucleobase chromophore. Much of the early work on the UV resonance Raman spectra of nucleotides and nucleosides was completed by the groups of Tsuboi, Spiro and Peticolas. 

Peticolas was the first to measure the UV resonance Raman spectrum and excitation profile (resonance Raman intensity as a function of excitation wavelength) of adenine monophosphate (AMP) [147, 148], The goal of this work, besides demonstrating the utility of UV resonance Raman spectroscopy, was to elucidate the excited electronic states responsible for enhancement of the various Raman vibrations. In this way, a preliminary determination of the excited-state structures and nature of each excited electronic state can be obtained. Although the excited-state structural dynamics could have been determined from this data, that analysis was not performed directly. 

What is remarkable is that all of these early measurements of the UV resonance Raman spectra of nucleic acid components involved computational and theoretical support to their experimental findings. For example, Spiro used CINDO calculations to determine the nature of the excited electronic states of the nucleotides [157], In the early and mid 1970 s, many researchers were also attempting to understand resonance Raman spectroscopy, the types of information it could provide, and a unifying theoretical framework to the intensities [147, 159-172], UV resonance Raman spectra provided some of the first experimental evidence to test the various theoretical models. Peticolas attempted to fit the observed experimental excitation profiles of AMP [156], UMP [151, 154] and CMP [152, 153] to the sum-over-states model for the resonance Raman cross-sections. From these simulations, they were able to obtain preliminary excited-state structural dynamics of the nucleobase chromophores of the nucleotides for UMP [151, 153, 158] and CMP [153], For AMP, the experimental excitation profiles were simulated with an A-term expression, but the excited-state structural changes were not obtained. Rather, the goal of that work was to identify the electronic transitions within the lowest-energy absorption band of adenine [156],... 

The major excited-state structural dynamics observed in UMP [173] were ascribed to modes at 1231, 783, 1680, 1396, and 1630 cm-1, in that order, while the major excited-state structural dynamics in the isolated chromophore occur along the 1231, 1680, 1396, and 1630 cm-1 modes in that order, with very minor contributions... 

A few of these UV resonance Raman studies have reported excitation profiles of oligonucleotides [158, 177], These studies show that the hypochromism in the resonance Raman intensities can be as large as 65% for bands enhanced by the ca. 260 nm absorption band for poly(dG-dC) and that the hypochromism can vary substantially between vibrational modes [177], In the duplex oligonucleotide poly(rA)-poly(rU) [158], similar hypochromism is seen. Although theUV resonance Raman excitation profiles of oligonucleotides have been measured, no excited-state structural dynamics have been extracted from them. 

Thus far, the only excited-state structural dynamics of oligonucleotides have come from time-resolved spectroscopy. Very recently, Schreier, et al. [182] have used ultrafast time-resolved infrared (IR) spectroscopy to directly measure the formation of the cyclobutyl photodimer in a (dT)18 oligonucleotide. They found that the formation of the photodimer occurs in 1 picosecond after ultraviolet excitation, consistent with the excited-state structural dynamics derived from the resonance Raman intensities. They conclude that the excited-state reaction is essentially barrierless, but only for those bases with the correct conformational alignment to form the photoproducts. They also conclude that the low quantum yields observed for the photodimer are simply the result of a ground-state population which consists of very few oligonucleotides in the correct alignment to form the photoproducts. 

Thus far, only one report of the UV resonance Raman excitation profiles of nucleic acids has appeared in the literature. The excitation profiles of calf thymus DNA [177] shows the same hypochromism as that observed in both single-stranded and duplex oligonucleotides. Also as expected, the excitation profiles are quite complex. Although an excitation profile is obtained for every vibrational mode, numerous bases are contributing to the Raman intensity observed in every vibration, each in its own microenvironment. Thus, the resonance Raman intensities currently are not useful for elucidating the excited-state structural dynamics of nucleic acids. 

The determination of excited-state structural dynamics in nucleic acids and their components is still in its infancy. Although progress has been made in understanding the excited-state structural dynamics of the nucleobases, primarily with UV resonance Raman spectroscopy, much work still remains to be done at that level to be able to extract the structural determinants of the excited-state structural dynamics and resulting photochemistry. Much less is known about the excited-state structural dynamics of nucleotides, oligonucleotides, and nucleic acids, but the static and time-resolved spectroscopic tools exist to be able to measure them. 

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