Elaine M. Marzluff

Low-energy collision-induced dissociation of deprotonated dinucleotides: determination of the energetically favored dissociation pathways and the relative acidities of the nucleic acid bases

Abstract Fourier transform ion cyclotron resonance mass spectroscopy has been used to examine the collision-induced dissociation pathways of all 16 of the possible deprotonated dinucleotides. These quasimolecular ions were generated by cesium ion bombardment of a mixture of triethanolamine, ammonium hydroxide and the dinucleotide. Collisional activation using continuous off-resonance excitation permits observation of energetically-favorable dissociation pathways. Dissociation products were examined over the range of center of mass energies from 0 to the minimum energy required to bring about complete dissociation of the reactant ion which did not exceed 7.7eV for deprotonated parent ions and 8.8eV for fragment ions in any of the systems. Semiempirical calculations were performed using the PM3 method, a variant of the AM1 method, to obtain gas-phase model structures and energies of the deprotonated dinucleotides and their collision-induced dissociation fragments. The acidities of the nucleic acid bases and dimethyl phosphate were calculated using the AM1 method. The deprotonated quasimolecular ions dissociate to yield several characteristic products. The major products formed in all systems are the deprotonated 5′-terminus base, the ion resulting from loss of the neutral 5′-terminus base, or the metaphosphate anion, PO−3. Insight into the relative stabilities of the fragment ions is gained by comparing the product distributions observed in each of the systems. The relative yields of products involving either the 3′- or 5′-end of the molecule suggest the 3′-terminus base is stabilized through hydrogen bonding interaction with the phosphate group. The relative strength of this stabilization follows the order guanine > thymine > cytosine > adenine. Additionally, the relative abundances of the deprotonated nucleic acid fragments suggest that the relative acidities of the nucleic acid bases follow the order adenine > thymine > guanine > cytosine. Only minor yields of sequence ions in which one of the phosphate diester linkages is cleaved are observed with these quasimolecular ions. Reaction mechanisms which account for the observed products are proposed.

Site-specific protonation directs low-energy dissociation pathways of dinucleotides in the gas phase☆

Abstract Fourier transform ion cyclotron resonance mass spectroscopy has been used to examine the low-energy collision-induced dissociation (CID) pathways of protonated dinucleotides. Collisional activation using continuous off-resonance excitation permits observation of energetically favorable dissociation pathways. Dissociation products were examined under multiple collision conditions over a range of average center-of-mass collision energies from 0 to 8.1 eV. Semiempirical calculations were performed using AM1 and PM3 methods to obtain gas-phase model structures of the protonated dinucleotides and their CID fragments. These calculations indicate that the proton is localized exclusively on one of the nucleic acid bases, with additional stabilization of some systems resulting from hydrogen bonding interactions between the bases. Protonated molecular ions dissociate to yield several characteristic products. The major fragmentation pathways are directed by the site of protonation leading to elimination of a protonated base, generally the 3′-terminus base. Exceptions are observed only in systems having thymine as the 3′-terminus base, where the major product is the protonated 5′-terminus base. These observations agree with the known relative proton affinities of the nucleic acid bases, and the existence of stable tautomeric structures of adenine, cytosine, and guanine which make these bases better leaving groups when protonated. In addition, application of statistical RRKM calculations to model the unimolecular dissociation dynamics of the reaction leading to the protonated 3′-terminus base provides an estimate of 1.9 eV for the activation energy associated with this major fragmentation pathway. In some systems, moderate yields of other fragment ions are also observed. Only minor yields of sequence ions are observed with these quasi-molecular ions. Reaction mechanisms accounting for the observed products are proposed.