Dissociation energies of deoxyribose nucleotide dimer anions measured using blackbody infrared radiative dissociation

Electrospray ionization (ESI) mass spectrometry (MS) is a powerful technique for obtaining molecular weights and structural information from biopolymers [1–5]. The application of mass spectrometry methods to the analysis of oligonucleotides has recently been reviewed [4, 5]. The base composition of DNA strands can be obtained from accurate mass measurements of double stranded DNA [6]. Sequence information can be obtained from both single and double stranded DNA using tandem mass spectrometry (MSn) [4, 5, 7–9]. Using several dissociation methods, McLafferty and co-workers obtained complete sequence information for a 50-mer nucleotide and nearly complete sequence information for a 100-mer [7]. Sequence construction algorithms have been used to determine the primary structure of oligonucleotides up to 20-mers from low energy collisional activation spectra [9]. In combination with solution-phase techniques, such as the Sanger method, MS and MSn have been used to verify sequences of DNA [4]. Mass spectrometry techniques have also been applied to the identification of antisense [10], labeled [11], and posttranscriptionally modified [12] oligonucleotides where other methods are often unsuitable. In ESI, gas-phase ions are formed directly from solution. This makes possible the direct coupling of mass spectrometry to solution-based separation and amplification techniques, such as HPLC or PCR. For example, Muddiman et al. demonstrated that sequence information can be obtained from oligonucleotides after amplification using PCR [13]. Noncovalent complexes are often observed in ESI mass spectra if source conditions are adjusted to minimize collisional activation in the electrospray interface. Many studies on the interaction of biomolecular complexes in the gas phase have been reported, including DNA–protein [14] and DNA–drug complexes [15] and DNA duplexes [16–19]. In the condensed phase, Watson–Crick hydrogen bonding between guanosine–cytosine and adenosine–thymidine pairs plays a fundamental role in the structure of DNA duplexes [20]. Recent results show that Watson–Crick base pairing can be retained for short duplexes in the gas phase (4- to 7-mers) [19]. This conclusion was based partially on dissociation activation energies for several complementary and non-complementary oligonucleotide dimers measured using blackbody infrared radiative dissociation (BIRD). For the complementary duplexes, the gas-phase activation energies were correlated to the binding energies in solution. The interactions between individual nucleobases have also been extensively studied in the condensed phase [20–24]. The binding energies of guanine–cytidine have been measured in chloroform and dimethyl sulfoxide using IR spectroscopy [20–23]. The structure in DCCl3 and deuterated dimethyl sulfoxide solution has been determined using NMR and in the solid phase using X-ray crystallography [20, 23]. In these nonaqueous solutions and in crystalline form, guanine and cytidine are Watson–Crick paired. Gas-phased dimer formation enthalpies of derivatized nucleobases have been measured. The energy for G · C is significantly greater than that of any of the other pairs [18]. Ab initio calculations have provided both binding energies and structures of guanine–cytidine pair in the absence of solvent [24–26]. However, the more rigorous calculations only include the nucleobase pairs and no phosphate and deoxyribose moieties. The binding energy of the Watson–Crick pairs determined from Moller–Plesset ab initio calculations are quite strong, 23.7 and 12.9 kcal/mol for G · C and A · T, respectively [24]. These values are in good agreement with those measured for derivatized nucleobases in vacuo [18]. Here, the dissociation kinetics and pathways of several nucleotide dimers are investigated. Threshold dissociation energetics are obtained using BIRD. From these values, information about the structure of these dimers is deduced. Molecular mechanics is used to obtain low energy structures for all nucleotides. The combination of both experiment and theory suggest that Watson–Crick hydrogen bonding for the guanosine–cytosine pair is retained in the gas phase.