Structural motifs of DNA complexes in the gas phase

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Abstract

DNA duplexes are known to be quite stable in the condensed phase but recent mass spectrometry results have shown that DNA complexes are also stable (at least for a limited time) in the gas phase. However, very little is known about the overall shape of the complexes in a solvent-free environment and what factors influence that shape. In this article, we present recent ion mobility and molecular modeling results that address some issues concerning the gas-phase conformations of DNA duplexes. Examples include the effect of metal ions on Watson–Crick base pairing, investigating the onset of helicity in duplexes as a function of strand length, comparison of the stability of C·G and A·T base pairs, and examining the formation of quadruplex structures.

Introduction

DNA duplexes are stabilized by a number of factors but hydrogen bonding between bases on the two strands and base stacking within each strand are the major contributors [1]. Solvent is believed to be just as crucial to the stability of the duplex because it can provide screening of the negatively charged phosphate backbones [2]. Thus, most structural and characterization studies of DNA are performed in the condensed phase and obtaining gas-phase data has been believed to be nearly impossible as an increase in charge repulsion from the absence of solvent screening and the reduced favorability of base stacking should significantly destabilize the duplex structure [2].

However, in 1993 Ganem et al. [3] and Light-Wahl et al. [4] demonstrated that DNA duplexes could be successfully transferred, intact, from solution to the gas phase using electrospray ionization mass spectrometry (ESI-MS) [5]. A year later, Doktycz et al. showed that DNA duplexes could survive in the gas phase for hundreds of milliseconds in a quadrupole ion trap [6]. Since that time, a number of papers and review articles have been written describing the usefulness of mass spectrometry in characterizing DNA duplexes in the gas phase [7], [8], [9], [10]. The types of results gathered from these studies range from the determination of base composition and sequence of DNA strands to the identification of ligand binding sites and post-translational modification sites. Despite this wealth of data, the overall gas-phase structures of the duplexes and the factors that influence these structures and the interactions between the two strands are not well understood (not to mention whether the gas-phase structures are representative of their solution phase counterparts or undergo major conformational changes).

Several groups have attempted to address these issues using selective dissociation of the duplexes and have shown that specific hydrogen bonding and base stacking may, indeed, be conserved in the transfer from solution to the gas phase. Gabelica and DePauw, for example, used collision-induced dissociation (CID) to examine a series of 16-mer duplexes [11], [12]. They observed that the relative kinetic stabilities of the gas-phase duplexes correlated well with relative stabilities in solution measured by thermal denaturation (monitored by UV spectrometry). CID fragmentation yields also paralleled calculated solution melting enthalpies. They attributed these results to the retention of hydrogen bonding and base stacking interactions in the gas phase that are present in solution. Schnier et al. studied the kinetics of dissociation of several complimentary and non-complimentary 4- to 7-mer duplexes using blackbody infrared radiative dissociation (BIRD) [13]. They observed that activation energies for dissociation of complimentary duplexes were higher than those of non-complimentary duplexes and these activation energies correlated with solution dimerization enthalpies, thus, indicating that Watson–Crick pairing was conserved in the gas phase (corroborated by molecular dynamics calculations). Griffey et al. examined the CID fragmentation of 8- to 14-mer duplexes with mismatched base pairs and observed preferential cleavage at the site of the mismatch (suggesting that Watson–Crick pairing was preserved in the rest of the duplex) [14].

Although these studies indicate that DNA duplexes conserve a portion of their solution phase character in the gas phase, their overall conformation in the gas phase remains a major question. In solution, DNA duplexes are often helical, taking on A-, B-, or Z-forms, but very little is known about whether these helices exist in the gas phase and what factors influence them. In this article we will report on some of our recent ion mobility and molecular modeling studies that have focused on these issues and examine factors such as how metal ions affect Watson–Crick pairing and the importance of strand length and sequence on the overall conformation of the duplex.

Section snippets

Ion mobility measurements

The mobility of an ion (K) is simply a measure of how fast the ion drifts through a buffer gas (νd) under the influence of a weak, uniform electric field (E) [15].νd=K⋅EFor large ions, the mobility also depends significantly on the geometric shape of the ion. Compact ions with small collision cross-sections undergo fewer collisions with the buffer gas and hence drift faster than more extended ions. Thus, Eq. (1) can be re-written astd=C1Ko=C2σwhere td is the drift time of the ion; Ko, the

Metal ions and Watson–Crick bonding

The importance of metal cations interacting with DNA was first realized in the 1920s when Hammarsten reported on the need for metal cations to be present in cells to help neutralize the overall negative charge on DNA [33]. However, metal-DNA studies did not really begin in earnest until the late 1960s after Rosenberg and co-workers discovered that cisplatin (cis-[Pt(NH3)2Cl2]) was an effective antitumor agent and ensuing work suggested that the binding of Pt to DNA bases was largely responsible

Conclusions

Although the results presented in this article are far from comprehensive and much work remains to be done, they do provide several important insights into the gas-phase conformations of DNA duplexes and higher-order complexes and some of the factors that affect these conformations:

  • (1)

    Watson–Crick pairing is enhanced in dinucleotides that are cationized by d10 metals.

  • (2)

    Duplexes can retain helical structures on the ms time scale in the gas phase. These helical structures are size dependent (first

Acknowledgement

The support of the National Science Foundation under grant CHE0140215 is gratefully acknowledged.

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